Science topics: MedicineImmunology
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Immunology - Science topic

Immunology is a broad branch of biomedical science that covers the study of all aspects of the immune system in all organisms. It deals with the physiological functioning of the immune system in states of both health and diseases; malfunctions of the immune system in immunological disorders (autoimmune diseases, hypersensitivities, immune deficiency, transplant rejection); the physical, chemical and physiological characteristics of the components of the immune system in vitro, in situ, and in vivo.
Questions related to Immunology
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 I am wonder if any one have a good experience in the IHC multiplex satin. i will study the co expression of biomarkers. any suggestion and recommendation please. 
thanks 
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I used too run chromogenic multiplex for her2+ER+PR, in If i did her2+ER+PR+ki67. All on the discovery platform.
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I am working on peripheral T-cell Lymphoma and looking for methods, assay how to see that isolated granulocytes are activated. I would be very grateful for suggestions.
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May be this article can help
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Hi all,
I am new to immunology field, need some help to distinguish CD3, CD4, CD8 and CD25. What are differences between them for T cells? How to decide which CDs are specific for T cells, B cells, neutrophils and macrophage.
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I think it will be best if you read a Basic Immunology book that has a chapter on hematopoiesis (Basic Immunology by ABul K Abbas and others) to understand why and how you can make the discriminations of different cell populations. Company catalogs will provide you information on the antibodies one can purchase from them, basic knowledge is in the books.
With best wishes,
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Hi,
Anyone here using FloJo to analyze data for instance lymphocytes population, please show how will be the outcome after analysis using this software?
In terms of statistics ?
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It's better if you provide more details about your questions. based on my experience, the flowjo software can show distinct populations based on their marker expression intensity. in a flowjo plot, one dot represent one cell, so you can also get the percentage of every population you have. these are basic functions that flowjo have, for more detailed, you can search on the official FLOWJO website or there are some useful youtube video that show how to analyse flow data by flowjo.
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I want to download transcriptomes from different types of immune cells T, B, Effector T cells, plasmatic cells, monocytes, macrophages, etc. It can be from microarray, RNAseq or scRNAseq from human tissues
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Hey guys,
I have a list of full mouse TCR sequences that I got from my vdj scRNA seq experiment, I am just wondering if there is a tool that might help me to predict what these TCR might recognize?
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Not really. There's been a ton of effort going into developing tools to do this but most are in early stage development and not ready for routine usage. The main issue is a lack of training data that can be used to refine these algorithms.
There are new in vitro high-throughput functional TCR antigen profiling techniques that have come online in previous years, including the one we developed: https://www.researchgate.net/publication/336310777_Rapid_selection_and_identification_of_functional_CD8_T_cell_epitopes_from_large_peptide-coding_libraries
These approaches are powerful because they let you directly test large libraries of candidate sequences in an unbiased way. If you have any interest in trying out our method, send me a DM and we can talk further.
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if possible please elaborate the phenomena also
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Progenitor cells in the tumor are usually Granzyme K+ and slowly proliferate. As the cells flip towards transitory/effector-like they become Granzyme B+ and stay Granzyme B+ as they further exhaust. So in essence there are Ki-67+ (with the capacity to proliferate) cytolytic cells and non-proliferating/terminally exhausted subsets.
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To provide some context:
Once an antigen is processed endogenously or exogenously by the APC's (ex: dendritic cells), it is then presented on MHCI or MHCII as a peptide.
specific naive Tcells have a matching Tcell receptor that is complimentary to the presented peptide. considering the incredible diversity of peptide antigens that can be presented, what governs the production of Tcells having the exact match to the presented peptide?
What is the mechanism of Tcell receptor diversity considering that somatic hypermutation is designated to Bcells?
(picture adapted from: Molecular biology of the cell - Garland science 2008)
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It is stochastic, or random. The receptor repertoire based on vdj, alpha beta v delta gamma and n and p additions result in a nearly unlimited. Umber of potential receptor rearrangement. The naive t cells enter secondary lymphoid organs at the instruction of chemokines and 4andomly test against antigen presented in the context of correct mhc. In this scenario, it I'd the antigen that selects the correct tcr, resulting in clone proliferation. The t cell is the passive player.
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Please let me have a simple question, "Should 7AAD be washed before Flow?". After I stained 1 million cells with antibodies and washed them, I will add 5uL of 7AAD. After incubation for 5 minutes in RT, should 7AAD be washed before the flow cytometer run? I mean, should the cells stained with 7AAD be washed 1-2 times before the flow cytometer run? Or, can I run the cell suspension that includes 7AAD as it is?
My 7AAD product does not have any protocol, and although I search on the internet, each protocol I found differs widely.
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Vipul Batra Thank you very much for your information!
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i am in my 6th semester and i am looking for some good colleges other than IIT's that are offering masters in immunology and molecular biology in india. can i get some recommendations on the same?
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Can I also get recommendations for universities offering masters in the same subject abroad as well. I am also looking for those options as well.
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I want to detect a specific protein in a pool of exosomes from plasma by ELISA, but I don't sure if the exosomes before ELISA test are o not disrupt using RIPA buffer or another methods.  What is the different? The capacity to bind at well is the same in one of this way?
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Hi André Cronemberger Andrade I read your comment and I would like to know if you have any protocol for what you mention about sensitizing the elisa plate to later detect surface proteins in exosomes. Do you use elisa plates of any particular brand?
I would appreciate it if you could help me, thanks in advance.
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Hello,
I am considering measuring total IgE, as well as anti-dsDNA IgE in mouse serum (most likely via ELISA) and I was wondering whether anyone has any experience in this. The animals in question are lupus-prone and non-immunized. Does anyone have recommendations regarding serum dilutions for such ELISAs? Regarding total IgE I have found recommendations in the range of 1:10 - 1:50 (for non-immunized animals). However, I couldn't find any information on anti-dsDNA IgE ELISAs. Maybe serum anti-dsDNA IgE concentrations are too low for ELISAs.
I would greatly appreciate any feedback!
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Do you already have reagents or a kit to measure total IgE and anti-dsDNA IgE? I would suspect the most straight forward way to measure total IgE is with a kit. Then for the anti-dsDNA IgE, you'll need to coat plates with dsDNA (followed by serum, then conjugated anti-IgE). Unfortunately, you'll probably just have to experiment to find the best dilution. But for anti-dsDNA IgE in an otherwise healthy mouse, I'd probably not dilute the sample at all and just report the result as O.D. Then test different dilutions depending on how you're setting up your standard.
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We are trying to build up a "Quick Step Guide for Young Scientists Playlist" to assist young researchers in their journey of scientific writing and publishing. Playlist link: https://www.youtube.com/playlist?list=PLwyWKnsS4EkgcQ1Wnkx7FIxUsLsLUp0gb
We hope it will help many of you.
Share it with your friends and colleagues. All the very best!! Channel link: https://bit.ly/Subscribe_Learn_SciTech
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Also these are good:
1- Writing Research Papers. Student's Book: From Essay to Research Paper
Dorothy E. Zemach, Daniel Broudy, Chris Valvona - 2013 - 128 pages
2- English for Writing Research Papers (English for Academic Research)
Part of: English for Academic Research (11 Books) | by Adrian Wallwork | Mar 3, 2016
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#MachineLearning Hello researchers, I was wondering that what are the most remarkable applications of machine learning algorithms in Biological sciences? My naive thoughts are: DeepMind by Google: Protein Structure predictions #Epitope predictions by DTU - Technical University of Denmark in Immunology Predicting tumor entities by DKFZ German Cancer Research Center for Central Nervous System. Please add to this list, looking forward for an interactive discussion. #structuralbiology #bioinformatics #research #immunology #cancerresearch
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https://www.nature.com/articles/s41592-021-01380-4 Deep Learning based approaches for protein structure prediction have sent shock waves through the structural biology community. We anticipate far-reaching and long-lasting impact.
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My experiment consists of a ex vivo stimulation experiment on isolated human monocytes.
Each biological replicate consists of the same isolate of human monocytes (derived from the same donor) and for each biological replicate there are three treatment conditions. The dependent variable for the experiment is secretion of a specific cytokine. In the analysis I am comparing the mean cytokine secretion of each group against every other group.
I am however unsure when comparing these means whether the data points for the three treatment conditions from each biological replicate should be considered paired or unpaired, as this will obviously influence the statistical test I use?
Thanks,
Gabriel
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As far as I understand, your data points/conditions are nested within individuals (donors) creating non-independence. In other words, the same individuals provide data for each condition. If that is true, then a paired test such as repeated measures ANOVA would be most appropriate. One way to examine this is by looking at whether the observations across conditions are correlated. For paired observations, you will typically find a positive correlation of the DV scores across conditions reflecting the non-independence of the data points.
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I'm a biomedical science graduate and hope to continue my MSc in immunology. I would like to know the career options I can have with master's in this discipline.Is it a good area of study. What are the MSc programs available related to immunology? please, recommend me.
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Immunology is a branch of biomedical science that deals with the study of immune system in health and disease. It has applications in various disciplines of medicine like oncology, virology, bacteriology, and organ transplant. With a Masters in immunology, you have various options open like Clinical Research Assistant in hospitals and clinics, technical support in clinical instrumentation like flow cytometry and ELISA-based techniques, and as a teaching faculty in universities.
Immunology is an interesting subject. If you wish and have a liking for immunology you can study further and complete your PhD in immunology and become an immunologist. This will be an added advantage as it will help you acquire a much higher position in academics, research or pharmaceutical industry.
All the Best.
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Egypt has the highest prevalence of HCV. What is the major risk factor or history behind it?
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Hello.
CIBERSORT or CIBERSORTx calculates "p-value" for each analyzed sample. This value seems to suggest the reliability of CIBERSORT analysis for each sample. If it will be p>=0.05, we may exclude the sample from the downstream analysis. However, sometimes we experience that more than half of our samples are excluded by the p-value. It will make the downstream analysis difficult.
Does anyone have a similar experience, and how do you handle this issue?
I am looking forward to your valuable advice.
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Sorry it is outside my aera of expertise
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Hello.
I would like to check cell changes by culturing T cells isolated from PBMC in amino acid-rich or deficient medium. How long should I culture in rich and deficient mediums to check cell changes?
It is also a concern whether it should be cultured in rich/deficient medium from the beginning, or whether it should be cultured in normal medium at first and then moved to culture them.
I'm sorry that my English is not good! Please give me a lot of answers! :)
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To obtain serum from fresh blood, it should be drained to a dry collecting tube. We have access to Buffy coats with CPD as the anticoagulant. How can we obtain serum from this?
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Let's change it this way, Its possible to get serum from EDTA-K3 plasma? the answer is yes.
Briefly, Centrifuge sample for remove of blood cells and keep buffy coat plasma at -20 °C. After thawing of the sample, dispense in another tube 1.0 ml of plasma add 26 μl of 1 M CaCl2 and incubate overnight in +4 °C. A clott is present at the end of incubation that must be squeezed and removed, obtaining clotting factors free serum.
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Antigens are coupled covalently to carboxylated magnetic beads, after immunologic reaction it is planned to elute antibodies and use the for next experiments. We have already tried to elute antibodies with low pH buffers like 50 mM glycine (pH 2.70) and 100 mM citrate (pH 3.0). Eluted antibodies were rapidly neutralized with 1M Tris buffer (pH 8.5), however, repeated run did not show positive reaction with eluted antibodies. Please, help find the best protocol!
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The difficulty is that if Antibodies bind nicely to your antigens it's usually that they are very affine to them and subsequently separating them is sometimes denaturing the antibodies themselves. They are some Mild elution buffer made up by Pierce you might give a try on that.
Other than that is to use maybe a higher salt content to protect antibodies but this is rarely working perfectly
Some more litterature you might want to read
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Respected researchers or professors,
I am Kumar Sharp, currently a third year MBBS student from India. I will be graduating medical school in 2024.
I have a very keen interest in Pharmacology, drug development, infectious diseases, microbiology and immunology. I have done my best to develop my interests in research and development, public speaking and leadership, publishing work in COVID-19 as well.,which you can see from my profile.
I would like to pursue my future career in these fields and teaching. I belong to a middle class family and do not have adequate resources to apply for international exams.
Can you all suggest if there are any ways to apply for these specialization in countries other than India.?
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China is the best choice, there are vast opportunities for the young
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Hi all,
I am trying to do a cell sort and collect equal numbers of CD3+CD8+ and CD3+CD4+ t cells from naive C57BL/6 mouse spleen. Theoretically 70-80% of CD3+ cells should be CD4+ and 20-30% should be CD8+, but when sorting the other day, the yield of CD3+CD8+ was only ~3% (we need at least 1 million, so this is not nearly enough), with 85% being CD3+CD4+ and the remaining likely be dendritic and NK cells. Based on the sort data, it is unlikely there was an issue with the antibody itself, nor the concentration used (I've also used it recently with no issues). Any thoughts on why I am experiencing significant loss of these CD3+CD8+ cells in comparison to the CD3+CD4+ t cell population?
Thanks for any advice you can provide!
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Bispecific antibodies, trispecifics or other formats have been and are still explored as powerful drugs against various cancer types. However, potentially these therapeutic agents could have a much broader application range. What are the reasons, researcher focus on cancer (beyond the obvious ones like incidence rate, death toll...)? The concept of bridging two or more epitopes on different cell types could in my mind potentially be extended to quite a vast range of targets. What about engaging bacterial cells? And what about non-peptide epitopes? Curious to hear some interesting thoughts!
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Hi,
Very important question. Moreover, the question is why (.... from my current knowledge please correct if I'm wrong) from more less 100 mAbs approved by FDA only 2 are bispecific ..... I think another trend is competitive for the "multispecificity" of mAbs. Its cocktails of mAbs (cocktails therapy). See some examples below:
1) atoltivimab + maftivimab + odesivimab-ebgn
2) bamlanivimab + etesevimab
3) IgG1(M777-16-3) + IgG2(62-71-3)
4) bamlanivimab + etesevimab
I think that cocktails therapy will be a more dynamic trend than "multispecificity" (because of higher and higher availability of different monospecific monoclonals)... and many other reasons,
Best regards
Tomasz
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I was wondering if anyone has any experience using alternatives for FCS in their HL60 cell culture (or similar cells), I've read that they can sometimes be cultured serum free. However long term I was wondering if there were any alternatives.
I'm happy to do a side by side comparison myself, but any starting input would be great.
Thanks for any help
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Fcs alternatives for HL 60 neutrophil like cell line is FPS (slightly different).
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Hi all, I did an IF staining for FoxP3 and CD3. Apparently, I can't do double staining due to the antibody available both from the same host. I just wonder if FoxP3+ cells must be overlap with CD3+ t cells? I saw more Foxp3+ cells than CD3+ cells in this case and wonder if the Foxp3+ I saw was a false positive. Any idea? Thanks in advance!
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FoxP3 + should be CD4+CD25+ Tregs. of Couse,FoxP3 should overlap with CD3+ T cells. usually CD3 should be expressed on the cell surface, while FoxP3 should be expressed in the nuclear of cells. they should co-expression in one cells. if you see they are differently localized. it means it is a unspecific binding and it is not correct. I often see some published paper they are wrong! confocal will detect them more accurately!
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Hello there, i'm working on a differente approach for Plaque Reduction Assays, and i would like to ask you, professional working with this type of technique: which are the major problems found by you when working with PRNT?
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Overall, PRNT has been reliable and a mainstay for virology and vaccine development. Here are the limitations I've found personally:
1) The throughput of the assay is somewhat limited due to the necessity of plaque counting as opposed to an automated read-out of reporter gene activity. I've really only done manual counts, and it looks to be somewhat difficult to automate the process. There are scanners with software for counting, but the visibility of the plaques and contrast with the uninfected monolayer can be limiting and the software may not give accurate counts. Related to this is the need to have the plaque size of the target virus being uniform so that counting is accurate.
One workaround is to examine the dilution series of each antibody and count only the two wells that bracket the endpoint, such as the 50% reduction in plaque counts. That can often be done in a hurry or to get preliminary data around the endpoint titer, but you lose a lot of information that way.
Another caveat for plaque counting either by human or machine is the necessity for cell monolayers to be uniform and complete enough in each well to get accurate plaque counts. If the cells do not form a 100% uniform monolayer by the end of the plaque growth against which the plaques can be counted, then it gets tricky, especially if the plaques/monolayers are stained with a dye rather than by immunochemical staining which may be better at showing the infected cells on a bad monolayer but is more time intensive.
2) The biosafety containment needs to be commensurate with the target virus biosafety. Often, researchers use the replicating virus of interest in the PRNT as opposed to a pseudovirus or chimeric virus which can be made to have lower biosafety requirements but will often have a reporter transgene if one is going to all the trouble of constructing these.
3) For some viruses that are more difficult to neutralize than others, it can be a little tricky to normalize the input virus between assays. This is more of a problem with the relationship between virus and neutralizing antibody concentrations, but practically speaking, inherent variability of the virus input can alter the results up or down depending on how much virus ended up in that assay. That can create more work in trying to put a large dataset together and have the assay data from each run match each other, say by the use of the results of a standard serum/antibody used in each assay.
4) One meta-problem that I've encountered is not the assay's fault, but rather the design of the assay with respect to the target cell. Viruses can enter the cells of various cell types in various tissues by different mechanisms, so the use of a non-physiological cell type for the neutralization assay can give nice, consistent results, but results that aren't useful going forward. For example, the human cytomegalovirus (HCMV) field for years employed primary human foreskin fibroblasts as the cells in the assay for which the neutralization of virus infection by antibodies was measured. This use is/was due to the very limited range of cell types that are available for permissivity of this virus for replication and plaque formation, so the cell type used is one of convenience (which itself is relative since these cells have limited passages). HCMV vaccine development experiments used these cells, and it wasn't for decades was it discovered that viral entry into endothelial cells is a more accurate assessment of neutralizing antibody responses compared to blocking entry/infection in a fibroblast. Thus, the fibroblast data wasn't particularly enlightening in the context of neutralization mediated protection. The cell type in the assay should be scrutinized when embarking on PRNT assays rather than treating them like another reagent in the assay.
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We are reaching you out because we are working on a project at the RWTH university Aachen, called Epi-Blood, which enables counting of leukocytes in frozen blood retrospectively, with high accuracy and requires only low volumes. www.Epi-Blood.de
We would like to understand the demand of the service that we will provide in the research sector. Therefore, we ask for just 2 minutes of your time to answer only 5 multiple choice questions. If you feel like contributing, please use the link below to start the survey. Thank you for your participation!
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I submitted a survey. Best of luck!
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Hello Everyone,
How to perform proliferation assay in terminally differentiated effector cells, I am not very successful getting proliferation in these cells. Please anyone suggest me to how to do it successful way or if anyone overcome similar situation share your knowledge that would be great help.
Thanks in advance for your time
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Live cell imaging with specific marker proteins
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Vertigo is a condition that can make it feel like you or your surroundings are spinning, sometimes leading to a loss of balance, according to the U.S. National Library of Medicine.
Coronavirus 2019 or COVID-19 is a novel entity which had led to many challenges among physicians due to its rapidly evolving nature. Vertigo or dizziness has recently been described as a clinical manifestation of COVID-19.
So, Are dizziness and vertigo COVID-19 Symptoms? and why?
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HI All,
I'm doing a survey as part of an Audacious program (https://www.startupdunedin.nz/audacious), which essentially is a StartUp initiative at Otago University. I'm curious to understand what level of programming do biologists these days need during their day to day research.
For all the biologists out there here are some questions to start the discussion on this topic:
1) Have you done any programming till date? If so which language did you use and for what purpose?
2) How have to overcome programming limitations? For example, did you get the work done through bioinformaticians, or sought help from your programming friend, etc?
3) Have you used online biological databases for your research? If so, which one?
4) How much of artificial intelligence have you used in your research? Do you see AI potential in your current work?
If you have anything else to add, please feel free.
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Please have look on our(Eminent Biosciences (EMBS)) collaborations.. and let me know if interested to associate with us
Our recent publications In collaborations with industries and academia in India and world wide.
Our Lab EMBS's Publication In collaboration with Universidad Tecnológica Metropolitana, Santiago, Chile. Publication Link: https://pubmed.ncbi.nlm.nih.gov/33397265/
Our Lab EMBS's Publication In collaboration with Moscow State University , Russia. Publication Link: https://pubmed.ncbi.nlm.nih.gov/32967475/
Our Lab EMBS's Publication In collaboration with Icahn Institute of Genomics and Multiscale Biology,, Mount Sinai Health System, Manhattan, NY, USA. Publication Link: https://www.ncbi.nlm.nih.gov/pubmed/29199918
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Sincerely,
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Eminent Biosciences.
Mob :+91 97522 95342
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How could immune changes due to allergen immunotherapy favor or be deleterious to patients with allergic diseases?
Would there be a difference during the build-up phase versus the maintenance phase of AIT?
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Dear Dr Larenas,
I invite you to read these articles to understand the relationship between SARS-CoV-2 and allergen immunotherapy:
Best regards,
Pr Hambaba
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*Sudanese Society of Clinical Immunology and Allergy (SSCI&A)*
💢 *Announcement* :
In collaboration with the Sudanese Junior Doctors Association SJDA, we are delighted to invite you to a highly scientific talk on COVID-19.
🌹We proudly announce the contribution of our renowned members:
🎤 *Dr Gehad Elghazali* (Consultant Clinical Immunologist, UAE),
🎤 *Dr Shuayb Alkhalifa* (Consultant Clinical Immunologist, UK),
🎤 *Dr Amal Hassan* (Consultant Paediatric Immunologist, Qatar),
🎤 *Dr Nahla Erwa* (Consultant Clinical Immunologist, Sudan)
SSCI&A Secretary General
🎤 *Dr Hadeil Morsi* (Immunology ST5, UK).
*We are expecting a great discussion on the use of convalescent plasma in the treatment of COVID 19 and better understanding of the immunology of the disease in general*.
Live Event will be hosted by this page.
Time : 20:00 Tuesday 26 May 2020
🇬🇧19:00 UK
🇸🇩20:00 Sudan
🇸🇦21:00 Saudi Arabia
🇦🇪22:00 UAE
Special thanks and gratitude go to *Dr . Abdalazeem Ibrahim* for the organization and coordination of the whole event.
Sudanese Society Of Clinical Immunology And Allergy *SSCI&A*
External relations office.
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Ashraful Hoque sir, please share your opinion on this...
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If there is Immunological Memory, there is also possibility of;
Immunological Dyslexia.
Immunological Dementia.
Immunological Amnesia.
Is it not ?
And such effects might occur as a result of adaptations in epigenetic system of individuals accompanied by ageing, co-morbidities, polypharmacy etc ?
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in order to select an antibody and co stain IHC slides
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Dear Nicolas,
I invite you to see this article:
Best regards,
Pr. Hambaba
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I'd like to have your opinion on this. What is the likelihood that a SARS-coV 2 variant emerges that can use antibody dependent enhancement?For example, if vaccine-induced antibodies against the S protein RBD become non neutralizing due to new mutations, or if their concentration becomes too low. Some interesting information in this preprint https://doi.org/10.1101/2021.02.22.432407
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Polyvalent vaccine commonly used in vaccination policy from different serotypes for the same disease.The occurance of vaccine Anti_BRD clonality is a lot and more narrow but if antibody enhancement in vaccinated region ,no clonality elsewhere to compasate,you must consider these variant invaccinatin programm under scientfic base vaccine must be strain variant vaccine(Include the variant in the region.
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I am currently working with Chlamydia infected HeLa cells. The protocol given to me by my PI states this should be happening within 2-10 min. However, he and I have given it a trial run and even after 45 min, we did not observe any lysis happening (under microscope). The basic run down of the protocol is to remove media, wash cells in with DI water, discard, then add ~2 mL of water per well (6 well plate) and observe under microscope. I am currently going through a couple more trials and am thinking of repeating the washing step and adding some form of mechanical shearing.
Is our estimated time too optimistic? Is there something I'm missing?
Thank you.
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Hi Samuel,
Even though this discussion is from 2016, I was wondering if you are able to comment on your observations of the Hela cell lysis? In particular, do you still see the membrane, just non-spherical? I am currently attempting a hypotonic lysis of HEK cells, but for the wash step am only adding water to a cell pellet and discarding the supernatant with out resuspension as I want to avoid premature lysis of cells before a final resuspension step in water and sample transfer. If, I can, I will attach an observation of HEK cells after 15 mins. The membrane is not circular and you can visibly see strands coming from the cell after staining with Propidium Iodide. I plan to run some proteomics experiments on the cells, so I need to be absolutely sure that they are lysed. Any help would be greatly appreciated!
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Hi there , i am consulting about tetanospasmin effects on immunological response without vaccine , what happens before the toxin enters to the CNS ? how the toxins enter to the CNS and PNS?
What is the response of the immune system against the bacteria and the Toxin separately ? Which are the receptors implicated on the initial inflammatory response ?
are there specific recognition sites that display very high affinity for a receptor?
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Cl.tetani spores after infection to wounds germinate due to the action of tetanolysine which favour anaerobe enviroment for getmination and production of tetanospasmin which resposible for stiffness of muscles mainly massetric muscles thigh.
Immunoprophylaxis of tetanus is restricted to toxin neutrlization.Tetanus toxoid is an aluminum hydroxide suspension is given for routine propgylaxis and a single injection will induce protective immunity in to 10 to 14 days
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By taking an example, spleen cells from mice-A are adoptively transferred to mice-B having RAG-/- and also mice-B is infected by a virus(eg: MHV).
  • Will there be any immunogenic reaction against this viral antigen in mice-B due to mediation by MHC of mice-A(non-self)?
  • What if the mice-B is RAG+/-?
  • Is there any possibility for alloreactivity along with this viral(against) immune response if mice-B is RAG+/+?
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@* My answer is no.
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Hi,
I want to make the IsoFlow Sheath liquide in our lab by following the chemical composition given by the company ( mentioned down below) , i want to know if any of you tried this before and if it works without any risk during the aquisition process. and i would like to know also if there are any tips or precautions to take when preparing the solution.
Chemical composition for 10L :
Sodium Chloride......................7.93 g/L
Disodium EDTA........................0.38 g/L
Potassium Chloride.................0.40 g/L
Monosodium Phosphate.........0.19 g/L
Disodium Phosphate..............1.95 g/L
Sodium Fluoride......................0.30 g/L
Kind regards.
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IsoFlow in Beckman coulter or sheath fluid in BD machine are the isotonic buffers solutions which may be replaced by Autoclaved and filter sterilized PBS.
You should be extra cautious while using in-house sheath buffer as any particle or growth may block your tubes in flowcytometer.
We have used Autoclaved and filter sterilized PBS in BD instrument and it never caused any issue.
Once you use any in-house buffer, the parent company will consider it void of warranty. So, never let the company people know that your are not using their isoflow.
Good luck,
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Hi,
I'm about to start a proliferation test for lymphocytes based on CFSE dye, but i don't have much knowledge in cell culture field. I would like to have some help from an expert on this field who has a valide protocol for that functional test.
Best regards.
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Please see my CFSE proliferation protocol attached and two papers for the results. The outline is for B cells, but it's easily adaptable to any other lymphocyte population.
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I want o start up my home-based Bioinformatic project on the immunological approach to post-covid complications.
or something related to probable drug discovery for covid complications.
Can anyone help me with ways or ideas in doing so?
If someone is already doing similar bioinformatic-based projects and need an apprentice, I am will to join as an apprentice.
I am a very hard working person please help.
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Is there any public patient RNAseq data repository related to COVID? If there is, and if they have FASTQs of them, any research may be done with those data by you at home. It may depend on the amount of meta-data connected to them, but you can search some relationship between some factors and some gene/gene set expressions.
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Bone marrow produce our immune cells & maintain homeostasis throughout the physiological system. Temperature, one of the factor among many, plays significant role in immunology immuno ontogenesis. I was trying to find the literature reporting the bone marrow temperature, but couldn't find any. If you find any of such literature, please share. Thank you
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Huggins, Blocksom and Nooman (1936), reported that the more centrally located bones of the extremities, the cranial bones and the sternum of Rats, Rabbits and Pigeon have temperatures similar to that of the peritoneal cavity. Whereas the peripheral bone marrow of the extremities showed significantly lower temperatures. (https://journals.physiology.org/doi/abs/10.1152/ajplegacy.1936.115.2.395)
However, in 1952, Nicholas Petrakis, published a paper on the determination of temperature of the human bone marrow in the Journal of Applied Physiology. The authors actually reported the temperature of the human bone marrow but the article is closed access so you have to pay to get it. here is the link https://journals.physiology.org/doi/abs/10.1152/jappl.1952.4.7.549?journalCode=jappl
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Usually if immune response to molecule A isn't elicited, then another known immunogenic molecule B is conjugated to A and delivered into the body.
The response is that the immune system elicits antibodies against molecule A, molecule B and molecule A-B conjugate.
Then in that case, do live vector vaccine delivery systems like Lactococcus/E.coli also get an immune response against them?
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I think yes.
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what's relation between IgA and IgG
mucosal vaccines is likely induce better sIgA than a needle vaccine. what about it stimulates serum antibody, such as IgG ? Is serum antibody correlate to sIgA? Who can tell me or has any reference answer my unknowing?
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Thanks
I am looking forward your publish.@Maral Barzegar-Amini
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when covid vaccine for children will be available in INDIA?
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What are the significant challenges and opportunities of the Computational Immunology (CI) as an emerging scientific area?
Where is the CI's tending?
What is the role of AI?
What challenges the simulation of immune system solve?
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I produced a chimeric IgG from a murine IgM. The IgM murine antibody could recognize the antigen in nature form (a receptor on the cell surface) by flow cytometry and by ELISA (in this case, the antigen is a recombinant protein). But the chimeric IgG is able to bind only the antigen (recombinant protein) by ELISA.
My question is: Did the isotype modification and chimerization process changes the affinity of my antibody?
How the constant chain could modify the affinity of an antibody?
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IgM is basically 5 IgGs glued together, which means that for the same affinity, you will nevertheless have substantially different avidity.
IgM is usually made first, and the individual Fab regions are usually pretty rubbish at binding antigens, but IgM has ten in total: the premise is that "rubbish x10" works out being quite good on aggregate. Getting ten crap binding sites in very close proximity can compensate for the fact that individually they're not very good. The body then takes whatever IgMs kinda work and hypermutates them to make (potentially much higher affinity) IgGs.
If you translate an IgM straight to an IgG context (without this secondary hypermutation), you lose the pentameric structure, and you're working on the base avidity of an IgG, which has only two binding sites. What works for IgM is not, consequently, guaranteed to work for an IgG.
So, short answer: affinity is likely unchanged. Avidity is markedly reduced.
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I'm using the BrdU antibody of the roche kit but i can't see any signal.
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In this case, it might be better to use another proliferation marker such as EDU because BrdU staining is mainly depending on denaturation by Hcl or you could try heating instead of Hcl treatment.
Good
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From your expertise, I want your consult how to get this projects since I have good experience on molecular techniques
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Even though i don't know how to get a project in whatsoever field but i think you can contact via various sources in order to know more about the same .I suggest you to get a project on role of B and T cells in fighting against Corona virus with special reference on enhancement of helper t- cells in B cell activation.
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The THP-1 cell showed the classical monocyte (CD14++ CD16-) behavior. If we want to study the non-classical CD14-Cd16++ without drawing blood and isolate the monocyte with FACS, what can we do?
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Dear all,
I haven't found a paper yet that claims THP1 is a classical monocyte. Would you know of one? Thank you!
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Despite diverse etiologies in erythroderma , the end stage immunologic pathways are similar. As results there are similar clinical findings irrespective of causes. But in some cases systemic corticosteroids are indicated. Is there any logic of not prescribing systemic corticosteroids in all cases of erythroderma?
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Dear Prof. Gheita, Md Abu Monsur Dinar, Hans-Joachim Kremer, thanks for your valued contribution. I appreciate your viewpoints.
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We already know answer would depend on the tissue gene expression profile, we also know it would depend on the particular HLA allele, it also depends on the affinity threshold to accept that there's a chance for binding ... I don't mind if the answer is in general terms or for a particular case (tissue, haplotype, allele...), but is there already any evidence to answer this question? Let's say 10%? 20%? 50%? Don't mind if the answer arises from biological studies or theoretical computer algorithms...
Much better if your answer is supported by a cited paper.
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think this idea is achieved at the biological level possible but very difficult and complex. But possible implementation by bioinformatics as well as via computer Algorithm. Good luck to do this .
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I am currently in the process of optimising a human monocyte-derived macrophage: naive T allogeneic cell co-culture. I am aiming to identify whether specific macrophage polarisation triggers naive T cell induction to Tregs. As the culture is allogeneic I will need some level of additional stimulation for T cell activation, in order to measure Treg induction by macrophages - we have already found that co-cultures with only macrophages and T cells and no additional stimuli, show negligible T cell activation. I am hoping that all that is required is T cell stimulation with anti-CD3, although I will try additional conditions with anti-CD28, as well as IL-2. The co-culture will last 3 days.
My main question is which specific approach is best to activate the T cells. I cannot use anti-CD3 dynabeads, as macrophages phagocytose them. Consequently, I was planning on using plate bound anti-CD3. However as the macrophages are matured from monocytes, they are already in the well and don't respond well to detachment and replating, and therefore I think using platebound will be difficult. Consequently:
-I could use soluble anti-CD3, however I have read that in order to actually crosslink CD3 for activation, plate bound is considerably more effective.
-I could possibly pre-activate the naive T cells (using dynabeads or plate bound Ab) and then transfer them to the macrophage wells? However, with this approach I am unsure how long I would need to incubate the T cells with stimuli for before transferring them.
-I could use non-bead based CD3 crosslinkers such as this one by Stem Cell: https://www.stemcell.com/immunocult-human-cd3-cd28-t-cell-activator.html . However, I have no experience with this product, so if anyone does and can share their experience that would be great.
If anyone has any experience or views on what they think the best approach to take would be, I would be very grateful to hear them.
Thanks.
Gabriel
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Many thanks for this excellent questions. These advance immunological information needs to more and extra to describe and explain this type of stimulation of cell mediated immunity ( T cells Immunity.
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I would like to stain a tissue for different markers, and as I there are not enough colours that I can use, I would like to do it sequentially. Thank you in advance!
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Hi janmaris@Janmaris Marin
you showed that you got good results of destaining the first antibody, I wonder if the striping buffer that you used was glycine-SDS(pH 2.0)in the Daniel Pirici article which your mentioned above.
Best wishes,
Lixin
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Hello everybody,
I am investigating the influence of a toxinon various clinical-chemical and haematological parameters in fish.
I have three treatments (control, low dose, high dose) in my experiment and 4 replicates (n=4). So a total of 12 tanks. Now, the fish from one tank are counting as dependent samples and therefore actually pseudo-replicates. Does it still make sense to sample more fish per tank? Do I have to pool the resulting data afterwards, as I statistically wouldnt be "allowed" to evaluate the fish from one tank on a per-fish basis? If we now assume that I sample 3 fish per tank, i.e. 12 per treatment (3x 4 replicates) and I have no significant tank effects, could I then increase my "n" to 12?
Regards
Finn
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Designed as a two way analysis of variance (treatment x tank with three replicates per tank) and the analysis showed no significant tank effect, you are effectively getting the same experiment as 12 replicates/treatment in a one way AOV..
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I encountered great difficulties in LDL oxidation, and I have no idea which step was wrong. Here’s my protocol:
The LDL was dialyzed in G2 Dialysis Cassettes (Molecular weight cutoff 7K) to remove the EDTA. Next, the LDL solution (5 mg/ml) was oxidation with CuSO4 (24hrs, 37°C, final concentration 25 µM)
To stop the oxidation, I add 1.25 mM EDTA, then dialyzed again to move out Cu2+ and EDTA. The final volume of the oxLDL was similar as the beginning.
Finally, I confirmed the oxLDL in BCA assay but found that its concentration was only 1 mg/ml. That mean I had lost 80 % of oxLDL. Any suggestion of these protocol ?
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It is difficult to find out the LDL oxidation process at 234 nm. You will get nice conformation by Thiobarbituric Acid-Reactive Substances (TBARS) assay.
  • Preparation of Oxidized LDL :-
  • Dissolve LDL (5 mg) in PBS (1 mL). Add PBS (1 mL) containing CuSO4 (10 μM). Transfer to 3.5 cm sterile plastic culture dish and incubate for up to 48 h at 37°C. Dialyze against 12 L PBS for 48 h at 4°C (two changes of 6 L each). Determine protein content by the Lowry method.
  • Measurement of LDL Thiobarbituric Acid-Reactive Substances (TBARS) in Oxidized LDL . Add LDL (50 μg) to NaCI (150 mM, 1.5 mL) in a 13×10 cm culture tube. Add TCA (20% w/v, 0.5 mL) and TBA reagent (0.5 mL). Boil at 95°C for 1 h. Cool with tap water. Add butan-1-ol (2.0 mL) and vortex mix vigorously. Centrifuge at 4,000 g for 15 min. Remove top layer and measure fluorescence (excitation at 515 nm, emission at 550 nm).
  • For more detail protocol is written in our recent published journal. See the link below.
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The unlighted part in treatment of COVID-19 is immunological role of pathogensis and how its effect ?? cytokine storms reduction necessary or not ?
New discussion
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Being a clinician with an on going project on arthritis and HCQ200MG as primary drug of chronic symptomatic arthritis, Immune Modulator is more helpful than any other Immune inhibitors.
Most of morbidity or mortality are because of immune component of COVID which can affect any organ and system.
Commercial guidelines for Medicine is most important challenge for medical professionals to be encountered
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Can someone conclude to me how we measure immune responses in Animal models in pre-clinical studies?
I know there are many ways and many essays. I want an opening to know how to start thinking in this part of immunology.
Thank you
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Hi (copy from your question into discussion loop ...)
If your aim is study immunogenicity of human drugs in animal models then best way is analysis of guidances by FDA/EMA :) and total ADA (anti drug antibodies), fraction of Nab's (neutralising - high affinity antibodies) and seroconversion time. Such data could be colected after sigle dose of human protein. Sometimes pilot study is useful. Sometimes best way to analyse immunogenicity of human drug in animal model is simply analysis of drug pharmacokinetics. Then you just see what the difference is with Nab's because this fraction of antibodies increases the clearance of the drug that is administered. You can measure the most important cytokines and cells. The meaning of such work increases when your model is relevant in pharmacokinetics and pharmacodynamic meaning. Please remember that using the animal models for comparision test versus reference drug immunogenicity could be valuable (see guidelines below) but translation into humans impossible and off value. Please remember that different doses of the duman drug can induce different immune responce. For example higher dose of the drug may give lower immune responce if drug has immunosupresive action. Moreover in case of some drugs multiple dosing may decrease immune responce for example ustekinumab or infliximab ....
Some explanations from old FDA guidance could be usefull in your work (document is not available now)
Animal immunogenicity assessments generally do not predict potential immunogenic responses to protein products in humans. However, when differences in manufacturing (e.g., impurities or excipients) between the proposed product and the reference product may result in differences in immunogenicity,
measurement of anti-protein antibody responses in animals may provide useful information relevant to patient safety. Additionally, significant differences in the immune response profile in inbred strains of mice, for example, may indicate that the proposed product and the reference product differ in one or more product attributes not captured by other analytical methods.”
GUIDANCE FOR INDUSTRY SCIENTIFIC CONSIDERATIONS IN DEMONSTRATING BIOSIMILARITY TO A REFERENCE PRODUCT FDA 2012
" A risk-based evaluation, focused on the mechanism of action of the therapeutic protein product as well as results of animal and in vitro evaluations should be performed to determine the need for collection of preand post-dose cytokine levels in the early phase of clinical development. In case of a clinical adverse event, such an evaluation may provide evidence to support the clinical diagnosis of cytokine release syndrome and help distinguish this entity from other acute drug reactions "
Guidance for Industry Immunogenicity Assessment for Therapeutic Protein Products
Immunogenicity in animal models is not predictive of immunogenicity in humans.
Assessment of immunogenicity in animals may be useful to interpret nonclinical toxicology and pharmacology data.
Immunogenicity in animal models may reveal potential antibody related toxicities that could be monitored in clinical trials.
May reveal immunogenicity differences between biosimilar and reference product
The immunogenicity of therapeutic proteins- what you don’t know can hurt YOU and the patient
" Therapeutic proteins show species differences in most cases. Thus, human proteins will be recognised as foreign proteins by animals. For this reason, the predictivity of non-clinical studies for evaluation of immunogenicity is considered low."
but .......
" the comparison of the antibody response to the reference product in an animal model may be part of the comparability exercise both for similar biological medicinal products "
" Qualitative or quantitative difference(s) of product-related variants (e.g. glycosylation patterns, charge variants) may affect biological functions of the biotechnology-derived protein and are expected to be evaluated by appropriate in vitro assays. These differences and impurities may have an effect on immunogenic potential and the potential to cause hypersensitivity. It is acknowledged that these effects are difficult to predict from animal studies and should be further assessed in clinical studies. "
Guideline on similar biological medicinal products containing biotechnology-derived proteins as active substance: non-clinical and clinical issues
Some nice papers about could be useful
Hope it's helpful
Best regards
Tomasz
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Dear professors,
I would like to discuss about immunology of COVID-19 vaccination as follows:
1. If you have received COVID-19 vaccination, which immunoglobulin (s) should be used as marker(s) for vaccination?
2. The presence of IgG represents vaccinated, isn't it ?
3. How to distinguish between the immunity from natural infection of SARS-CoV-2 and vaccination ?
Thank you very much.
Dr. Methee
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Now, in a new, non-peer-reviewed study published in the journal bioRxiv, researchers show that this decrease is related to the disappearance of a family of antibodies called immunoglobulin M, or IgM, in blood plasma. In other words, IgM antibodies play a major role in neutralizing the virus and are part of the arsenal that the immune system uses to fight
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I would like to share with you this ambiguous case, herby clinical and biological resaults :
Gender : F
Age : 12 years old
Clinical case : - ENT infections / Buccal candidiasis / Anemia / Adenopathy / Splenomegaly .
- Viral serologies : Negative
- Vaccination : all done successfully
Biology : Immunoglobulins : IgG : 94.315 g/l , IgA : 0.826 g/l , IgM : 5.552 g/l
FC : CD3+ : 1151 c/µl
CD4+ : 766 c/µl
CD8+ : 25 c/µl
CD19+ : 164 c/µl
C56 : 52 c/µl
HLA DR : On Lymphocytes : 12 % / On Monocytes : 100%
Serique proteins electropherosis : Alb :23.69 %
Apha1 : 2.02%
Alpha2 : 4.86%
Beta : 6.54%
Gamma : 62.89 %
Immunofixation resauts :
Presence of multiple monoclonal components with low concentrations of various isotypes, against a background of very intense polyclonal increase in gammaglobulins.
We passing hard time to intrepret this resault , i would like to know your thoughts about this case.
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I just spoke to a rheumatologist about the case and she considered off the bat
Sjögren’s Syndrome
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I am wondering if anyone has had success with in vitro polarization (via cytokines) of Jurkat cells. My goal is to understand the affect of a drug on Treg induction and differentiation. I understand that this would be reasonable using human/mouse primary T cells, however this may not be practical and or feasible in my case, at least for now. I would like to first explore the effect of this drug on T cell activation/differentiation using Jurkat cells. I am having trouble finding literature on this topic. Thanks in advance.
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Hi Ben, here is an earlier paper from 2012 on the topic:
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Help! This mystery population (lookin' cool, in all black) is driving me crazy. I can't find this subset described anywhere.
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Have a look when convenient
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I have conjugated glycol chitosan with bolton hunters reagent, the FTIR spectra of glycol chitosan and glycol chitosan plus bolton hunters reagent look exactly the same. I need xy data to look into detail any slight difference between the two
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Hi, I'm running an assay measuring phagocyotosis and ROS production in PBMC's. I'm using Dihydrorhodamine 123 (Thermo) to measure ROS. I then stain and fix with PFA afterwards.
I've so far read conflicting things on whether you would be able to keep your cells fixed with DHR intracellulary, or would the results be less reliable? Any insight is much appreciated!
Mari
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Hi Mari,
Relative to ROS production: working with fresh human whole blood (not PBMCs), we found considerable DHR fluorescence following PMA stimulation in monocyte and neutrophil populations by flow cytometry. Anything between 20 - 30mM DHR123 in DMSO for a final cell-stain dilution of (1:20) was ok.
Stimulation time matters as well. A quick look at literature reveals that ROS generation is a fast process; so you may get good signal with PMA in whole blood anywhere starting from 30mins at RT. We noticed that the fold change in neutrophil oxidative index with PMA began to drop significantly after 2h of stimulation. Hence you want to stay within that range if using PMA.
With respect to other stimulants: these affect the fluorescence intensity as well. In my hands, Mtb bacteria whole cell lysate produced a relatively higher signal than the unstimulated control under conditions stated above but significantly less than that induced with PMA.
NB: We didn't try fixing cells for later acquisition; we read immediately after the staining process.
Best wishes,
Caleb
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Virology, microbiology, immunology, vaccine
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Please take a look at this useful link.
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I studied biochemistry and molecular biology in my bachelors and Immunology in my masters. I never came across a suitable course for general signaling pathways and principles. My interest is particularly focused on immunology but I would like to have a general overview just as much. Any recommendations?
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Both excellent choices but can add
Molecular Biology of the Cell in your shelf for some amazing reading
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My lab have some skin biopsies that were previously conserved in paraffine , and i wanted to know if it is possible to extract DNA or RNA from them.
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check the following link
it my be helpful
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The clinical treatment of COVID-19 infected individuals.
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I hope this
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Can anyone please suggest some related topic or area in microbiology and immunology except COVID/SAARS?
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molecular virology
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Hello,
I am reading on vaccines in general and I wonder if the beneficial effects of vaccines (as vaccination is the most effective choice when pathogens are to be stopped and prevented) lower the Darwinian fitness of immune cells such as T-cells and B-cells.
Thank you for your answers!
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may be
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Here what CDC writes about COVID-19 mRNA: " COVID-19 mRNA vaccines are given in the upper arm muscle. Once the instructions (mRNA) are inside the muscle cells, the cells use them to make the protein piece. After the protein piece is made, the cell breaks down the instructions and gets rid of them.
Next, the cell displays the protein piece on its surface. Our immune systems recognize that the protein doesn’t belong there and begin building an immune response and making antibodies, like what happens in natural infection against COVID-19."
1) From classic immunology we know that muscle cells are not the antigen-presenting cells.
2) We know that intracellular proteins are being presented in class I MHC to CD8 T cells, which in this case, have not been "trained" in thymus for this antigen.
So how are we sure, that these mRNA will not trigger myolytic /autoimmune response towards the muscular MHC class I/COVID-19 complex?
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