Melanoma - Science topic
A malignant neoplasm derived from cells that are capable of forming melanin, which may occur in the skin of any part of the body, in the eye, or, rarely, in the mucous membranes of the genitalia, anus, oral cavity, or other sites. It occurs mostly in adults and may originate de novo or from a pigmented nevus or malignant lentigo. Melanomas frequently metastasize widely, and the regional lymph nodes, liver, lungs, and brain are likely to be involved. The incidence of malignant skin melanomas is rising rapidly in all parts of the world. (Stedman, 25th ed; from Rook et al., Textbook of Dermatology, 4th ed, p2445)
Questions related to Melanoma
Most protocols do not include details about preparing cell lines for tumor inoculation and various labs seem to do things differently based on my personal experience. Could anyone please share what they personally do for the following things:
1. Do you keep the cancer cells warm or on ice prior to inoculation? Why?
2. What gage needle do you use (for subcutaneous)? Why?
3. Do you load the needle any special way to avoid shearing?
Thank you for sharing! Any other general related comments are also welcome. We are specifically working with various melanoma cell lines.
I am working on B16F10 melanoma cells. Yesterday I thawed the cells that I had freezed last week and saw some floating black spots. Is it a sign of contamination? If so, what type of contamination is it? This cell line produces melatonin black pigment too so I am not sure if it is normal to have black debris. This is passage 3 and I bought this cell line from ATCC.
Sorry to bother in this Sunday
But I would like to ask a few questions:
Does anyone know about how to induce the generation of polyploid giant cancer cells (PGCCs) in melanomas with "targeted drugs"?
How to measure the presence of polyploidy in the tumor?
How to induce the generation of giant polyploid cancer cells (PGCCs) in melanomas?
Is there any protocol to induce the generation of polyploid giant cancer cells (PGCCs) in melanomas?
Is there any way to study the immortalization and regeneration of tumors with cellular and molecular biology? Is there any protocol?
Matheus Correia Casotti
We isolated CD8+PD1+ T cells from the peripheral blood of a metastatic melanoma patient, with 99.5% isolation purity and 98% PD1 expression. We activated cells with Human TransACT "Miltenyi Biotec". After the activation phase had been finished, we left the cells in the expansion phase for 14 days, and then we performed a flow cytometry experiment to identify the phenotypic features of the expanded population. We found out that PD1 expression decreased from 97% to nearly 50%, but we still have CD8+PD1+ purity of 96%.
The part we didn't understand, what was the cause that led CD8+PD1+ T cells to decrease PD1 expression?
...For research purposes to help others. If you are or know of anyone who is who would be willing to share his/her story, I would greatly appreciate the introduction! Thank you so much!!
Dear fellow researchers,
my histo team and I came across some trouble in reducing background autofluorescent noise in FFPE tissue from primary melanoma. So far, we have been able to quench distinct portions of noise in 488 and 546 with Sudan-B. Still, we suffer from problems in the 647 channel and in 750/790 regarding some antibodies.
Can you suggest other reliably techniques, especially for primary melanoma tissue?
Best wishes and take care,
If any of you have worked with the MM28 cell line (uveal melanoma metastasis) how did you freeze it to maintain the best cell viability?
I have ordered A375 melanoma cell lines and the ATCC website recommended using DMEM+10% FBS. I was wondering if the cells can be grown in RPMI 1640+10% FBS as I am using it for some other cell lines.
Dear colleagues, we need to induce a melanoma-derived tumor in the monkeys. Whereas, murine melanoma cell lines are available, i can't find any similar monkey cell line. Is it possible and reasonable to induce tumors in monkeys by using human cell lines?
Can anyone please share Skin cancer(melanoma) dataset please.
I am using the B16-F10 cell line as a model for melanoma in mice. after treatment, i see an increase in the ratio of M1/M2 macrophages in tumor tissue. i suspect that the treatment I use either reprograms M2 TAMs to M1 or increases M1 migration into tumors from other sites? I was thinking of using a drug (after the tumor reaches 100mm3 in size) to inhibit the recruitment of any new microphage into the tumor thus I can distinguish whether TAMs already in the tumor are reprogrammed to M1 or M1 are migrating from other sites to the tumor.
I recently received these cells as a gift from one of my colleagues. I wish to use these in my research which is on anticancer activity of some cyanobacteria species. I couldn't find any broad information about these cells on internet or from literature. I am wondering if these cells are suitable for this kind of studies?
I am working on a deep learning-based analysis to detect Melanoma Cancer. I want to use all types of algorithms to get the best possible solution.
I need a melanoma dataset where the lines are classified. that is, the images are marked that they have lines and what category these lines belong to.
I’m interested in calibrating 2‐Deoxy‐D‐glucose in different concentrations in melanoma cell line. How do you recommend me examining the effect of 2‐Deoxy‐D‐glucose on the cells other than measuring the lactate levels? Do you recommend any genes that are affected in their expression levels to measure using real time PCR for example?
I have noticed morphological changes in primary melanoma cell cultures after switching to a different brand of FBS, the cells are circled in blue. Some cells began to form a more round shape with black particles attached to and surrounding the cells.
After the removal of antibiotics, the black particles seemed to increase as free floating particles with no particular motion. I assumed that it was mycoplasma contamination and tested them accordingly via pcr. The results were negative. I then assumed that it was potentially apoptotic bodies from low quality FBS so I decontaminated everything and purchased new FBS. After activating a stock, the cells seemed to be much healthier but after a few passages, have begun to exhibit the same changes again.
The cell passage number is <10 and I have passaged this cell line above 10 before and didn't see these changes so I assume its not straying further into abnormality. Once they are above 50% confluency, they morphological changes are gone and they assume their shape. Is this a potential other form of contamination or something that is exhibited by some primary cancer cell lines?
I am currently started working on the analysis of Melanoma Cancer by using DL Algorithms. I got to know that Metastasis is the spread of cancer from one organ to another. On the other hand, the physiological process through which new blood vessels develop from pre-existing vessels formed during the vasculogenesis process is known as angiogenesis. I need a proper explanation and relation between angiogenesis and metastasis.
The sun’s UV rays help your body make this nutrient, which is important for your bones, blood cells, and immune system. It also helps you take in and use certain minerals, like calcium and phosphorus. And while most people get enough vitamin D from food, children who don’t can get rickets, which softens and weakens their bones.
Moderate amounts of sun over your lifetime, especially in your teen and young adult years, might make you less likely to have problems seeing things at a distance (nearsightedness). But too much direct sunlight can hurt your eyes. It can lead to blurred vision and raise your chances of cataracts.
This also raises your chances of skin cancer. If you do it before age 35, you’re 60% more likely to get melanoma, the most serious form. Even one session can raise your odds of melanoma by 20% and other types by as much as 65%. If you want that all-over body tan, tanning lotions might be an option. Most are safe, but they usually don’t have sunscreen in them, so don’t forget to put that on as well.
Figure 3.1. TBX3and CERS1 are inversely correlated in melanoma: (A) The Cancer Genome Atlas (TCGA) melanoma samples (black line) were scored and ranked according to the Verfaillie invasive gene signature (Verfaillie et al., 2015). Vertical grey lines indicate expression of the indicated gene in each melanoma sample. The purple line indicates the moving average of (left panel) TBX3 or (right panel) CERS1 mRNA expression across 20 melanomas. Bioinformatic analysis using TCGA SKM. A correlation between TBX3 mRNA and melanoma invasiveness was performed in melanoma patient samples n=303.
We are trying to isolate DNA/RNA from thin primary FFPE melanomas. We use a protocol based on silica columns for the isolation of DNA/RNA from the same specimens.
Although the DNA that we get seems to work fine on subsequent enzymatic reactions (PCR, sequencing), we encounter inhibitory problems with the RNA samples. In particular, it seems that RNAs are contaminated with melanin. The amount of RNA that we have is very limited, 30 μl with a concentration of 20-50 ng/μl.
Has anybody used the CTAB-urea method or a column based method for removal of melanin from RNA preparations? Our problem is the very limited amount of the starting RNA.
Thanks a lot,
I am growing a Melanoma cell line expressing Ovalbumin (OVA) protein. I am looking for maintaining the stable expression of the protein by cell line and the researcher use G418 with a variable concentration in different papers. Can anyone have experience with starting the selection process? what is optimum concentration and how to select it? ED50?
Thanking you in advance.
I met a problem when I was doing frozen section for mice melanoma tissue. Based on the confocal results I got as attached, seems like the cell structures are destroyed (looks like a network) and all cells are CD8 positive (T cell markers) and F4/80 positive (macrophage markers). Is there any potential reason for this? I used 10% formalin to fix the tissue overnight. Is there anything wrong with this fixation procedure?
are there any murine cancer cell lines out there that once injected subcu will metastasize to the liver? I know there is LLC and some melanoma cell lines that will go to the lung. Appreciate any input!
actually I searched the related web sites, but the quantity of samples are maximum 20 or 30 pictures, but I saw in articles that there are datasets from theses 2 sources with enough number of samples (more than 100).!!!
Hello people of ResearchGate,
I need your help in identifying a contamination that has appeared in our cell cultures. We have discovered these contaminations in various cell cultures cultured in the same cell lab. When observed through a microscope, the contaminants appear as spherical objects, visible even with a 20x objective. The medium does not change colour, and when measured I have seen that the contaminants are all approximately of the same size with a diameter of ~9 mikrometer. My guess is that it might be some sort of fungi, maybe yeast, since they are so big and we use cell culture medium with antibiotics.
I have attached a picture taken with our microscope (20x objective) of the culture. It is supposed to be an adherent melanoma cell line (the big, darker stuff), and the contaminants are the bright, spherical objects floating above the melanoma cells. Does anyone of you have any idea about what it might be?
I am looking for a dataset of melanoma patient attributes. So for example age, sex, history of melanoma, history of cancer, family history, benign or malignant.
I hope my question was clear. Any help would be greatly appreciated.
I was asked to do a presentation on the following paper:
Juratli, Mazen & Menyaev, Yulian & Sarimollaoglu, Mustafa & Siegel, Eric & Nedosekin, Dmitry & Suen, James & Melerzanov, Alexander & Juratli, Tareq & Galanzha, Ekaterina & Zharov, Vladimir. (2016). Real-Time Label-Free Embolus Detection Using In Vivo Photoacoustic Flow Cytometry. PloS one. 11. e0156269. 10.1371/journal.pone.0156269.
In the introduction, it says something like this: "However, application of the PA technique to the real-time monitoring of embolus dynamics during a medical intervention in large vessels is challenging. The main goal in this paper is to demonstrate that the in vivo PAFC can rapidly detect emboli triggered by melanoma and (...)"
- My interpretation of this is that, for some reason, PA isn't enough so it must be used together with flow cytometry. I would like to know why and what is the difference.
Could any scientific researcher guide me to obtain SW480 or Human melanoma cell lines in Egypt. Thanks in advance
I need a dermoscopic image database to test an algorithm for automatic diagnose. In particular I am interested in images with blue-black colors within the lesion.
Can anyone tell me where to find it?
I have 92-1 of melanoma cell line , when i thawed them they were totally normal , but after first passage , the cells begun not growing well and not totally attaching on the bottom of the flask .
looking for your reply
Thank you very much
I am currently doing some CRISPR/Cas9 KO on human melanoma cell lines. I found in different articles the puromycin concentration for selection in A375 (1 μg/ml). But, I cannot find this information for the WM3211 melanoma cell line.
Has anyone ever use this WM3211 cell line for gene KO experiment?
In addition to clinical examination, dermoscopy and RCM help in diagnosing AHM. Which would be a better option between the two? Why?
We are working in our lab about melanoma. In each in vivo experiment, we digest tumors to have a single cell suspension. The problem is that the precntage of viable cells is very low, between 10 and 20%. We are looking for another protocol through which we can get a higher percentage of viable cells.
A prospective study has been published by Wen-Qing Li (JAMA Intern Med. doi:10.1001/jamainternmed.2014.594, online April 7, 2014) raising an intriguing question, stating that Sildenafil use for ED showed a statistically significant elevated risk of melanoma. We´re really concerned about this. Any insight regarding this important subject?
Hi all - I apologize if this has been covered before.
What can I expect from a tumor grown from B16-F10 cells in a C57B6/J mouse?
30 days after subcutaneous injection of 10^6 cells, I harvested a massive growth from the abdomen that I assume was a metastasis. However it was WAY bigger than the subcutaneous growth. It is also very soft/mushy.
Is that normal? Do these cells result in very soft tumors usually? The subcutaneous portion was much firmer.
Could it be that they grew too fast and the whole thing became necrotic? Unfortunately the mouse died the morning of the planned sacrifice, so I was unable to properly perfuse and harvest the way I wanted to.
I am doing research on melanoma detection for dark skin people. so it's very difficult to identify the dark skin images, please tell me the idea to get the dark skins.
My current job is make spheroids from melanoma cell lines. But i dont know how to identify spheroids. How can i know that i have a spheroid and not a cell cluster?
Please help me, this is my essay for the thesis
im trying to make spheroids from melanoma cell culture, most specific with cells A375, G361 and SK-MEL-1. If someone try please let me know how do you meke, because im new in this area. Thanks
Hi there I experienced problems with autofluorescence of RBC on FFPE sections of human melanoma. Used AF555 sek. Ab to detect. Was thinking about using the trueview form Vector labs but they suggest a certain mounting medium. I have to use ProLong Glas antifade mounting medium from ThermoFischer which has a higher refractive index since I'm working with an LSM microscope. Does anyone has experience already with Trueview in combination with mounting media others than suggested by Vector labs? Cheers, David
I am working on melanoma B16-F10 cell line and after treatment whit different concentration of drug I do SRB protocol then I read their absorb at wavelengths of 545 nm. now, how to analyze these results? What should be the range of control absorbency?
I want to inject B16 cells into C57BL/6 mice as a melanoma model to study tumor immunology. But I'm confused the injection type. Both subcutaneous and intradermal injection have been used. Which one should I choose?
Our B16F10 cell line from ATCC encountered massive cell death after switching the growth media from RPMI to DMEM.
We were stuck with RPMI( 10% FBS, 5ml P/S) for the first 2 passages with the lab mistakenly ordered DMEM w/o sodium pyruvate & l-glutamine. The B16F10 cells looked healthy as we split them at the subcultivation rate of 1:5 and passaged them every 3-4 days.
Then, the growth media was replaced with DMEM when the glutiMAX arrived. 5mL of glutamax (5mM) + 10% FBS, 5mL p/s was added to prepare the DMEM. The cells were split 4 days ago with the new DMEM and at a much lower ratio of 1:12. We checked the cells today and they were all dead. Shown in the pictures.
Any advice for the troubleshoot would be very much appreciated! Thank you in advice for the help!
I tested 25 ug/ml and 40ug/ml of Fluorescently labeled transferrin in melanome cells during different times of observation, but no fluorescent signal was detected inside the cells.
Is there anyone here who has had the same problem?
In my research, I want to treat melanoma cell lines with some inhibitors. Then I want to measure Braf protein activity. I want to do this with ELISA method. But I can't see any paper measuring Braf acitivity with ELISA. So I want to ask is it wrong to measure Braf activity with ELISA? and Can you send me papers related to ''measurig Braf activity with ELISA'' if you know??
In patients affected by metastatic melanoma (BRAF mutated) that were previously treated with target terapy (dabrafenib 75mg x 3 / day; trametinib 2mg / day) is it recommended to be treated with combined immunotherapy (Ipilimumab + Nivolumab) or there are better alternatives?
I’ve used the same normalization and chip to search a gene expression in R2 Genomics Analysis and Visualization Plataform, but I have compared very different cells (like lung cancer and melanoma). So my doubt is: is it correct/safe to say that a gene is more expressed in lung cancer, for example, than in melanoma (p<0,05) based on comparations made in R2 Genomics?
Ps: my intention is to prove this experimentally, and the results obtainned on R2 would serve as a guide for me.
I could not found the cell sizes of some cancer types such as; lung (NCI-H2126, ATCC CCL-256), breast (UACC-1179, ATCC CRL-3127), over (EFO-27, DSMZ ACC-191), melanoma (A-375, ATCC CRL-1619), pancreatic cancers (PA-TU-8988S, DSMZ ACC-204).
Could you share your information?
Thank you for your help.
I have just started to work with animal cell culture studies. I am working on certain novel tyrosinase inhibitors as skin whitening agents. Almost all the literature survey I have done is using B16F10 cells for inhibition assays. I have SK-MEL 31 cell lines which is also melanoma cell line but from human source. I didnt come across with papers using SK-MEL lines for tyrosinase or melanogenesis inhibition. I wonder if I can use SK-MEL 31 for the same experiments.
I tried to thaw several stocks from the -80 freezer that date back to one year and others from liquid nitrogen that date back to two years but they die continuously. I thaw them in 6-cm culture dishes but they are very weak and very slow. I waited for so long until one plate became confluent and then splitted it in two 10-cm plates. When they were passaged they became even weaker and every day I look at them, I find a lot of floating cells and nothing improves when I change the medium. N.B. I culture them in DMEM with 10% FBS.
I have used the common ABCD features for melanoma classification. However I want to explore some more suitable features. Could anyone help me?
Papers related to Melanoma Skin cancer using Image processing and the latest methodology being Incorporated to identify the Cancerous cells?
We are doing a anti tumour activity by expressing a gene through AAB DJ8 (Cellbiolabs).
Weirdly we see anti tumour activity in even in control AAV GFP vectors! We are using the Melanoma xenograft model and the AAVs are injected via the tail vein.
We don't purify the vectors as such; we have extracted the viruses using freeze thaw in presence of benzonase. Further we pass the extract through 0.45 filters and its resuspended in PBS.
Pls. free to ask me more questions if needed.
I’ve just made results of immunocytochemistry with B16F10 melanoma cell (1st antibody: anti-Sox10/ 2nd antibody: Alexa 488 green) and then I got total 7 images which are observed by optical microsope, DAPI, Alexa 488green, DAPI+Alexa488, optical+DAPI, optical+Alexa, optical+DAPI+Alexa.
However, i’m not sure which image I should put on my paper. Is there anyone who can help choose typical result images for appropriate result of immunocytochemistry with nucleous protein??
Dear colleagues !
I found the ISIC dataset to test my segmentation method for the skin lesion. However, I cannot find the corresponding ground truth. If someone worked on this dataset, please let me know where can I get their ground truth?
Many thanks !
I am working with the B16-F10 murine melanoma cell line in C57BL6 mice. I have seen several different growth patterns after s.c. injection. Usually the tumors develop as small black dots and progress into round black tumors. I have also seen spreading black patches that don't develop into raised tumors. Recently I have also seen brown scab-like growth after injection. Cell prep and injection was the same in all cases; can someone explain the differences or has anyone seen similar growth patterns?
Any advice would be greatly appreciated!
Dear Colleagues and Friends
I have faced a strange results in my previous experiment.
I had Melanoma cells in culture and after 4 days, I wondered when I saw that the melanoma cells were morphologically changed to a neuron like cells or more likely to melanoblast cells.
The most exciting part of this morphological changes, that prevent me to ignore, was that their growth have been stopped or significantly decreased without applying any specific/new chemicals.
So, I would like to know about any non-specific reason behind this phenomena.
Considering that, this morphological and metabolic changes were happened for a malignant/invasive type of skin cancer cells, the answer may results in a way to step forward in cancer treatment.
I am currently looking for a working unconjugated antibody directed against human CD39 (ENTPD1) for immunohistochemical analyses on FFPE tissue (Melanoma).
Thank you for any recommendations.
Actually we tried to develop a xenograft metastatic model of melanoma with NOD-SCID mice after ic injection of A375 metastatic melanoma cell line. After 4 wks by ic injection we only found oligometastatic disease, prevalently spread to bones or muscles.
Are NOD-SCID mice suitable for this model?
Many other groups used Nude mice for the same purpose, however with different cell lines. We will soon repeat experiments with other cell lines.
Does anyone have experience with both NOD-SCID and Nude mice xenograft models of metastatic melanoma?
I want to find out if skin tatoo has any effect in skin cancer
I need a commercially available or patient-derived melanoma in situ cell line for my research at MSKCC. Can anyone help me find one?
I am looking for a method to stain spleen sections to identify germinal centers in mice. What stains, antibodies, or protocols do you use to verify the identity of a germinal center? Thank you!!
I try to get primary cells from my melanoma mouse. But I failed several times (cell contamination or cell not growing). So is there anyone do this experiment before? would u please tell me your steps or relevant papas. Thanks
Cancer starts to occur with the disorderly change of healthy cells, which forms a mass known as a tumor. A tumor is of two types, Benign and Malignant. Malignant meaning that the tumor can grow and increase to other parts from the body whereas benign can grow but will not spread. The skin is the largest organ within the body, protecting against injuries and infections. Malignant melanoma is a very dangerous form of skin cancer with the largest number of incidents around the world. Early diagnosis quickly followed by its removal for malignant melanoma is very important for a patient to survive.
Melanomas and sun related skin damages occur exponentially in the last decades. Primary care providers are the perfect professional group to screen the skin of patients.
I've been looking for weeks finding only the same papers and none of these actually state the medium they used.
I need this for my master thesis and I would be eternally grateful if someone could help me out here.
I am planning to do a very preliminary experiment using a few representative solid cancer cell lines. Probably Three for each cancer types.
For Breast cancer, I am thinking to use MDA-231, MCF7 and maybe one HER2+ cell line. Since my research before is all about hematological malignancies, I would like to ask the experts' opinions about which cell lines are the representative or most commonly-used ones for the aforementioned three cancer types. Thanks
Approximately 30% of cancer patients respond to ICB therapy, some for very extended periods. ICB is an enormous step forwar for many cancers such as melanoma but still, the majority of patients do not respond or relapse after initial response. The discussion on this topi is hot and there is evidence for different causes of resistance. I would like to know your experience.
Im running some experiments on melanoma cells and betulin.
Can you tell me please which solvent I can use to solubilized the betulin?
Hi. I'm trying to transfect my Malme3M and A375 melanoma cell lines with using the HiPerfect transfection reagent. However, I'm having trouble transfecting these cells. I use the AllStar Cell Death positive control from Qiagen to assess the efficiency of the transfection (the more dead cells, the higher the efficiency). However, the cells just won't die. I used siRNA concentrations ranging from 7.5 nM all the way up to 50 nM. I used 3 or 4.5 µL transfection reagent. To form complexes I incubate for 10min in 100 µL Optimem medium before adding. I use 20.000 cells in a 24 well plate.
Does anyone have an idea what could be the problem? Should I use a higher concentration? Or add more transfection reagent?
We need a mouse melanoma cell line from from this genetically engineered model mTyr::CreER; BrafCA/+; Ptenlox5/lox5 ( please see link to paper)
i have scanned the ATCC, Wistar databases, emailed (the understandably very busy) authors of the paper . No success
I am having trouble with staining with B16 melanoma brain tumor samples using FoxP3 antibody on frozen samples, I am not able to see the positive cells I am using DAB staining followed counter stained with H&E. Please anybody have suggestions how to stain with frozen brain sections protocols.
Protocol to induce resistance to BRaf treatment
I want to do IP and roughly want to number of cells needed for decent amount of protein.