Cancer Animal Models

Cancer Animal Models

  • Hamed Karimian added an answer:
    Can anyone assist with benign and malignant tumor animal models?

    Hello everyone,

    I would like to create two animal models: benign vs. malignant tumor (with same tissue origin) by chemical compounds. Any input and protocol would be appreciated. 

    Hamed Karimian · University of Malaya

    look for (DMBA)

  • Jenny Woof added an answer:
    How different are the functions of IgG2a and IgG2c?

    there is hardly any info on effector functions for this isotype present in some mouse strains.

    Jenny Woof · University of Dundee

    I think you are confusing the names of the IgG subclasses in mice and rats. In mice, the subclasses are IgG1, IgG2a, IgG2b and IgG3. Rats have four subclasses known as IgG1, IgG2a, IgG2b and IgG2c. There is no direct correspondence between the subclasses in the two species.

  • Mark Jason Sinnamon added an answer:
    Does anyone know of or use a metastatic colorectal cancer model?

    I currently use the APCmin mouse model. However, this model only develops benign polyps in the intestinal tract. I am interested in continuing my studies in a colorectal cancer mouse model which develops metastatic tumours. I know previous papers bred APC with SMAD3 mice. However, this will take time to inbreed through 12 generations and unfortunately do not have time to do that.

    Thank you,

    Mark Jason Sinnamon · Massachusetts General Hospital

    Breedge,

    Another option is to obtain the cre-lox AKP mice created by Tyler Jacks' group.  I believe they are available through jackson lab at this point.  These mice have apc 9truncation), p53 (truncation) and kras (oncogenic lox-stop-lox) inducible upon adenocre exposure.  Alternatively, there is are AB and ABP mice with oncogenic BRAF under lox-stop-lox control.  Ken Hung developed a protocol to induce colonic tumors using these mice that do occasionally spontaneously metastasize to liver and lung.  I generated a handful of tumor cell cultures from these mice (in mixed c57bl/6 background) that are capable of colonizing the liver upon splenic implantation.

  • Hien Thi My Nguyen added an answer:
    Is anyone using TOV21G cells to induce tumorigenesis in balb/c nude mice using xenograft model?

    Hi,

    I'm conducting xenograft model using balb/c nude mice. I am using TOV21G cells to induce tumorigenesis. Would you please suggest how many cell numbers I should use? Which mouse condition? Which brand?

    I want to see the tumor growth.

    Thank you. 

    Hien Thi My Nguyen · Chung-Ang University

    Hi, thank you for your advice!

  • Safia Ezzine added an answer:
    Problem with 4T1 Tumor Model Development- Can anyone help?
    We have been trying to develop 4T1 breast tumor model in balb/c mice. We use 10^6 cells/100 uL/mice by injecting in the nape region subcutaneously. The cells grow vigorously upto 28 days post injection forming a large 3-4 cm long mass with a distinct 3 dimensional morphology growth under the skin (upon excision the mass weighed approximately 7 grams!!) . By this time period, there develops a break in the skin and a black scar like tissue formation occurs. The tumor then slowly starts regressing after day 28.

    Is it normal for such a large tumor mass to ultimately regress? Has anyone experienced the formation of a black scar tissue once the tumor develops? Is this black scar like tissue leading to tumor regression and if so, how can it be avoided?
    Safia Ezzine · French National Centre for Scientific Research

    According to my experience,  don't try to transfect this 4T1 breast cancer tumor! ONLY viral vectors works . in vitro it's really hard to transfect...and as Clemens said, The hypoxia means no bioluminescent probes ...but with Fluorescence you loose the (photons collected, so the amplification of signal=quantitative ). avoid GFP, IRFP....Good luck guys!!

  • Pieter R Roelfsema added an answer:
    How can we make a better effort at public outreach about the necessity of animal (especially NHP) models for understanding the brain?

    I thought the opinion article written by Pieter Roelfsema and Stefan Treue did a great job of explaining the complex issues involved in NHP research, as well as some of the research breakthroughs that have come as a result of NHP research, with positive consequences for brain understanding and human health. The article ends by encouraging researchers to engage in more public outreach to ensure continuing support. But the question of how this could be accomplished is not addressed.

    Pieter R Roelfsema · Netherlands Institute for Neuroscience

    Dear Andre,

    Thanks for pointing this out. There is indeed an increasing awareness among neuroscientists that it is beneficial to become more open about the benefits of our research. We should inform the public about our research to gain support and to combat misinformation.

    A useful resource - presumably known to all of us - is www.brainfacts.org, and there are other comparable websites. You can also work together with organizations such as "Speaking of research", and talk to the press when you uncover something new and exciting. I can recommend the paper on this topic by Holder (2014).

    All the best,

    Pieter Roelfsema

    Holdert, T (2014) Standing up for science: The antivivisection movement and how to stand up to it. EMBO Rep. Jun;15(6):625-30

  • Rangarajan Narasimhan added an answer:
    How to induce neuroblastoma?
    How can we induce neuroblastoma in rats
    Rangarajan Narasimhan · Sri Sankara Arts and Science College

    I am working with glioma. i have induced glioma by injecting c6 cell lines in the rat's brain

  • Santosh Kumar Yadav added an answer:
    Do you have any useful tips on how to weigh a laboratory rat?

    This may be a simple question, however, I face this problem every day. As you aware, there is a need to analyze the weight of the animal during study period. While weighing in an electronic scale, the rats are moving and the number in the digital screen keeps on changing. I used to record a number (which I observed frequently) from the screen as weight. 

    I don't have an idea on how to deal with this practical problem. Please share with me any useful tips to weigh a laboratory rat with accuracy.

    Thanking you in advance for your answer.

    Santosh Kumar Yadav · Central Drug Research Institute

    Thomas J R Beveridge have detailed the method which is in regular practice. Observe the readings for 2-3 sec and record the value which appears more times during up and downs.

  • Hien Thi My Nguyen added an answer:
    Reverse effect between in vivo and in vitro. What is the possible reason?

    Hi all,

    I constructed stable cell lines with my interested gene. It has inhibition effect on cell growth in vitro. However, when I use these stable cells for in vivo study, both SC and IP model, I got the reverse result on tumor growth compared to my in viro result. My gene enhaced tumor growth in vivo. I'm confused. Which can be reason for my result? Please suggest me some hypothesis. Or you can share me some tips when working with animal.

    Thank you so much.

    Hien Thi My Nguyen · Chung-Ang University

    Dear all guys,

    Thank you so much for your suggestions. I'll discuss it with my supervisor. I've never tried 3D culture before, so I have no idea on it. I performed tumor growth in vivo with and without matrigel. I realized that without Matrigel, my gene enhanced tumor growth  about 80% of tumor numbers while it only enhanced tumor growth about 50% number of tumors with matrigel. In details, when I used matrigel to conduct SC model, tumors were palpable after 2 weeks of injection but most tumors disappeared 1 week later and then appeared and grew again. I used TOV-21G cells, 5*10^6-10^7 cells/injection, cells prepared in PBS, balbc nude mice.

    My supervisor wants me to perform cancer animal model again to confirm if my gene really enhanced tumor growth in vivo before we find answer for this reverse effect. I'm a newbie to this kind of experiment, I hope someone can suggest me a protocol or some tips for animal model.   

    Thanks.

  • Krishnaa Mahbubani added an answer:
    What is the best In Vitro assay? Alamar Blue, Presto Blue, XTT or MTT?
    The Assay for in vitro activity..
    Krishnaa Mahbubani · University of Cambridge

    I have tried a variety of assays in an attempt to find a consistent and not overly time consuming method to get consistent data. What I have found with Alamar Blue, is while it is cheap and extremely easy to use (owing to it's low toxicity), it does need to be optimised specific to your cell line. If you cells are suspension lines, this makes it additionally a bit more complicated to optimise the time line. 

    I have a preference for MTT/MTS assay. They are not particularly time consuming and can be run in 96-well plates and in larger or smaller volumes dependant on sample size and repeatability options.  This can often be coupled with an LDH assay to give you a secondary set of tests for confirmation. 

    To overcome some of the challenges with inconsistencies as well as in terms of having one or two highly metabolic cells in a well over proliferating cells, I normalise my data using a bicinchoninic acid assay (BCA) to measure protein content within the cells being tested. 

    Like everyone has stated already, it is a very good rule to use two or three different assay methods for cytotoxicity. 

    Good  luck!

  • Lovett Evan Reddick, PhD added an answer:
    Has anyone used a cumate-inducible gene expression in vivo in nude or scid mice injected with tumor cells transfected with a cumate switch vector?
    I need to know if you can induce gene expression by injecting it into the mouse. There are plenty of examples of using the system in cell culture but I wasn't able to find anything about using it in animals. Thank you.
    Lovett Evan Reddick, PhD · University of Texas Southwestern Medical Center

    Recently, I've been in touch with SBI. I'll include the email correspondence below: 

    Hi Evan,

    Thank you for the background information. It helps to know your experimental goals and design in order to provide information that better addresses your needs.

    I would like to schedule a call with my colleague, Travis Antes, next week to discuss the in vivo performance of the inducible system further. As mentioned in the previous correspondence from Jake and Young, the test results show two key attributes for this system: tight expression with essentially no leakage and an ability to cross the blood brain barrier.

    Please let me know the best day and time to schedule a call (Tuesday through Friday) next week, after Travis returns from his business trip.

    I look forward to your reply.

    Cheers,
    Laurie
    Laurie Muscatel Goldman
    Sales Manager
    System Biosciences (SBI)
    265 North Whisman Road
    Mountain View, CA 94043
    Cell: 925.323.0569
    Office: 650.968.2200 x103
    Fax: 650.968.2277
    LGoldman@systembio.com
    www.systembio.com
    Visit of our Linked-in Page by clicking <image001.jpg>



    From: Evan Reddick [mailto:Evan.Reddick@UTSouthwestern.edu]
    Sent: Thursday, August 14, 2014 2:46 PM
    To: Jake Lesnik
    Cc: Travis Antes; Laurie Goldman; Young Kim
    Subject: Re: Contact Us Submission from Evan Reddick


    Hi Jake, Young, Travis, and Laurie-

    That sounds great, I’ll look forward to talking to Travis in the near future.

    A bit of background on what I want to do, so as you guys can better meet/anticipate my needs. I have a protein toxin that kills and/or growth arrests a particular cancer cell type. I would like to have an inducible lentiviral system such that I can stably integrate my toxin gene into the cell line, pick single clones using FACS and puromycin, and then introduce them into a mouse xenograft. Because the toxin so potently kills/growth arrests this cell type, leaky expression is a bad thing for me. After it’s been introduced and forms tumors, then I can IP or feed mice with the inducer, in this case, cumate.

    Thanks very much for the assistance, and I look forward to speaking more about your products. The main reason I’m interested here is that the data suggests that the cumate system seems to be less leaky than the Tet/on Dox inducible system that is available from several other providers.

    Cheers,
    Evan


    On Aug 14, 2014, at 1:48 PM, Jake Lesnik <JLesnik@systembio.com> wrote:


    Hi Evan –

    Thank you for your inquiry. In fact, we are pleased to inform you that we have recently developed and validated a water-soluble version of cumate that should help you do exactly what you are suggesting. See attached data from a similar type of experimental setup.

    I don’t have the exact cumate administration protocol, but I will ask one of my colleagues to follow up with this. Our lead scientist for this program, Travis, is on a business trip and may not be back in the office until Monday, but we will follow up with more information as soon as possible.

    Sincerely,
    Jake

    Jake Lesnik
    Director of Business Development & Marketing
    System Biosciences (SBI)
    265 North Whisman Rd.
    Mountain View, CA 94043
    Phone 973-986-5875
    FAX 650-968-2277
    jlesnik@systembio.com
    http://www.systembio.com



    From: evan.reddick@utsouthwestern.edu [mailto:evan.reddick@utsouthwestern.edu]
    Sent: Thursday, August 14, 2014 8:22 AM
    To: Travis Antes; Jake Lesnik; Young Kim
    Subject: Contact Us Submission from Evan Reddick

    A promotion was received from:

    Name: Evan Reddick
    Title: postdoctoral researcher
    Email: evan.reddick@utsouthwestern.edu
    Phone: 214.633.1373
    Institution: Univ. Texas Southwestern Medical Center
    Comments: I'm interested in xenografting tumor cells with stable integrations of a cumate-inducible gene into nude mice. Has this been done in the past? Can you provide me with any information regarding the administration of cumate to mice? Thank you in advance.

  • Guozhong Qin added an answer:
    What is the method to develop hepatocellular carcinoma in rat or mice models?
    I have to work on hepatocellular carcinoma in rats or mice using any chemical method.
    Guozhong Qin · EMD Serono

    Aflatoxin B1(AFB1) also induce HCC, but it induce hcc slower than DEN.

  • Kirsti Rouvinen-Watt added an answer:
    Are there any organisms that don't get cancer?

    Are there any multicellular life forms that don't get cancer or where cancer is very rare?

    What can we learn from these cases?

    Kirsti Rouvinen-Watt · Dalhousie University

    It has been suggested by Tian et al. 2013 (Nature 499: 346-349) that extremely high molecular mass hyaluronic acid is the reason why naked mole rats do not get cancer.

  • Frank Cannizzo added an answer:
    Does anybody know about cell line injections in mice?
    I am interested in developing a cancer model. I am using HEY and UCI (ovarian cancer cells). Mice are balb/c. Appreciate any suggestions.
    Frank Cannizzo · University Health Network

    You could take a look at the MM14.Ov cell line from ATCC (ATCC # CRL 6383). Depending on what you are looking for this might save you from having to culture your own. Good luck!

  • Maurizio Pea added an answer:
    What does Peto's paradox really mean? Are mouse tumor studies relevant?

    Peto's paradox states: "There appears to be no correlation between body size, longevity and cancer across species and the absence of such a relationship is referred to as Peto’s Paradox".

    Example: whales and elephants are massive cell conglomerates with long lifespans. Why don't they have high cancer rates, higher than humans? Why do humans not have higher cancer rates than smaller mammals? Is there a paradox? If so, what does it mean for the use of small, short-lived mammals such as mice as cancer models?

    Maurizio Pea · University of Verona and ULSS 22 Bussolengo, Italy

    I do not know if what I wrote in another thread may interest you.

    Staying within the carcinomas ( epithelial malignancies ), these are very rare in some anatomical sites (and when present, they are in the context of some syndromic complex), epididymis , seminal vesicles , uterine tubes , the small intestine etc. Conversely locations where cancers arise most frequently are the transition areas where active chronic inflammatory processes result in a high cells turnover (sometimes with metaplastic processes ) . In these circumstances carcinogens ( chemical and biological ) have more effect ( the bronchial mucosa, the cervix where there is a transition between squamous and columnar epithelium , the gastric mucosa which changes and becomes absorbent type ( intestinal metaplasia ), etc. . Other areas of transition are particularly subject to turnover following the sequence of hormonal secretory and proliferative stimuli ( ductulus in the breast , at the junction between the duct and lobule where the majority of breast cancers occurs , the endometrium etc). Probably this is not the sole cause of the difference in the incidence of cancer in various epithelia, but it can be one of the main switchboards. Seems to me that, for example, the seminal vesicles have been studied to find some intrinsic characteristic to justify their inability to develop cancer , but I think with few results.

    In other circumstances specific genes are likely involved. The keratoacanthoma of the skin that arises from structures of the hair. It shares many morphological and molecular characters with squamous cell carcinoma of the epidermis, however, are probably still active genes that regulate hair growth (anagen and catagen). Thus, this tumor grows and regresses on schedule by the hair cycle.

  • Denise Priolli added an answer:
    Why are the treatments initiated when the average tumor volume reached 100 mm3 in xenograft tumor model?

    In some experiments for investigate drug effects, we are generally using mice which have 100 mm3 tumor volume. This was a general process within ethical rules. I wonder how was this general process determined? Is there have any reason for using 100 mm3 tumor volume when it's starting therapy? Any reference is appreciated.

    Denise Priolli · Universidade São Francisco

    The size of 100 is good to heterotopic xenograft in cancer animal model, but in ortothopic xenografts, some time, 100 is no possible to obtain. How it will be possible 100 at colon, stomach, prostate, ovary cancer models, specially if the research is done with Balb c nu/nu, a very little murine? Other consideration is the body % area, size of tumor and the drug test concentration into the tumor. Is it possible reach the sufficient concentration in the tumor center with big size tumor? So, there are questions no answer yet.

  • Gudrun Lang added an answer:
    Can tumor markers (alpha fetoprotein, carcinoembryonic antigen) be performed on mice tissue homogenates instead of serum?

    Can tumor markers (alpha fetoprotein, carcinoembryonic antigen) be performed on mice tissue homogenates instead of serum? What are the other tumor markers which can be carried out on tissue homogenates of a HCC model of Swiss Albino mice? 

    Gudrun Lang · Allgemeines Krankenhaus Linz

    Wouldn't it be more informative to do a IHC stain for the same tumormarkers on the specimens? Or is it a matter of quantification?

  • How do I accurately measure tumor volume?

    I am using a DMBA-TPA carcinogenesis model and would like some input on how to measure tumor volume.  Some of my tumors (> 1mm) are not raised very high above skin so it is difficult to measure height?

    Thanks in advance

    Wolfgang Hohenforst-Schmidt · Coburg Clinic aff. University of Wuerzburg

    Hi Matt,

    this is the formula of Monga: (L*W*H)/2 which is doing pretty well for clinical issues.

    Monga SP, Wadleigh R, Sharma A, et al. Intratumoral therapy of cisplatin/epinephrine injectable gel for palliation in patients with obstructive esophageal cancer. Am J Clin Oncol. 2000;23(4):386–392.

    I totally agree with Joseph Ciccolini - they are all correct wrong. I do not agree with R.A. Laine as this measurement is much more time dependent after cutting out than expected (unless it is frozen but then the volume is wrong again) - and the same tumour consists of less water parts with progressing growth due to progressing central density wich is well known by the radius dependent IFP (Interstitial Fluid Pressure).

    Kind regards,

    Wolfgang

  • Fakhrul Khan added an answer:
    How to induce solid tumors in nude mice using leukemic cells along with matrigel?

    Is matrigel useful in induction of solid tumors in case of leukemic cells? or leukemic cells alone are good enough to induce tumor? Could anyone please suggest me the procedure for induction of solid tumor using leukemic cells if any? 

    Fakhrul Khan · Galgotias University

    http://ftp.sccwrp.org/pub/download/DOCUMENTS/AnnualReports/1987AnnualReport/AR87_70.pdf

  • Miles Mckenna added an answer:
    What mouse lung cancer and normal cell line are available?
    Do you know any normal mouse lung cell lines other than MLE 12 (which transformes Tumorogenic)?
    Miles Mckenna · Colorado State University

    What sort of lung cancer are you looking for? There are non-small cell lung cancers (NSCLC) as well as small cell lung cancer (SCLC), NSCLC includes adenocarcinomas, suamous cell carcinomas as well as a few additional NSCLC's. I added a link to the ATCC website for lung cancer and normal cell lines for all organisms, including mouse. 

  • Deanna Larson added an answer:
    Does anyone have knowledge on genotyping for APCmin mice?

    Hi All,

    Does anyone have a good protocol for genotyping APCmin mice? I have the primers from the paper below, but was wondering if anyone has recommendations for the PCR setup (i.e. annealing temp, cycle number and so on). Thanks much. 

    http://cancerres.aacrjournals.org/content/65/22/10623.long#sec-2

    Deanna Larson · Utah State University

    This is what we used.... Hope it helps.  We used amplitaq Gold MM.

  • Serhiy Souchelnytskyi added an answer:
    Who knows a study that characterizes the tumors in patients with unexpected long-term survival?
    Sometimes we see patients living much longer than expected. Despite the advantages of multimodality treatment, their prognosis seems to be "hopeless" (e.g. gastric cancer with peritoneal carcinomatosis and metachronous liver mets). But they are able to live free of tumors for years. This raises the question: why?
    Serhiy Souchelnytskyi · Karolinska Institutet

    Thanks for taking up this issue! I have also experience of patients who are alive despite very bad prognosis.

    Studies showing diagnostic problems (like the Finish study) expose managerial reasons of unusual survival.

    Studies showing biological reasons have been difficult to find, at least for me.

    When I do tests for individualization of anticancer treatment, I see huge differences in responses of different patients/tumors/tumor cells, even if the histological diagnostic is similar. If to take into account that we may expect more than 100 molecular profiles of a single type of cancer, e.g. IDC for breast cancer, individual differences in molecular profiles of tumors are certainly among key reasons.

    I am sure that individualization of molecular profiles of tumors (genomic, RNAs and especially proteins) will lead to more publications of unusual survival...

    With best regards,

    Serhiy Souchelnytskyi

  • Annamari Zainana added an answer:
    Hind limb ischemia in SCID mice?
    We are facing a problem in induction of hind limb ischemia in SCID mice. We tried common femoral artery ligation (only knot) and also cut, but it is not inducing ischemia as expected. The limb is totally functional, no difference in the muscle mass and the histo-pathology also showed negative results.

    Interesting thing here is, when we tried the same surgical methods in BALB C mice, which is the background strain for the SCID mice, the surgery worked perfectly well and the limb lost its function completely. Histo-pathology results were also positive in BALB mice.

    So can anyone suggest the best surgical method for inducing hind limb ischemia in SCID mice? Please cite any references.
    Annamari Zainana · Charles River Laboratories International, Inc

    Hi!

    Did you ever try to perform the proximal ligation site higher up (more proximal), in the area of iliac artery, and when performing the distal ligatures closing all side branches and/or closing the femoral vein together with the artery? These procedures increas all the injury severity. It has been shown that  occlusion of the femoral vein together with fem.artery very probably results in the loss of the injured limb, which is usually not the desired result, as the level of ischemic severity usually is aimed to be severe but curable. Obstruction of the vein is often not recommended due to that, and the fact that in human arterial diseases, the ischemia is based on occlusion of an artery, not vein.

    Also, dissecting the vessel(s) between the ligatures produces more grave injury since a single cut has been shown to be less effective due to fast growth of collaterals after the cut (You can find excellent information and advice for the mouse hindlimb ischemia from e.g. jove.com)

    I'm sure all this you already know, but I have noticed that the ischemia severity can be increased by simply closing and dissecting more circulatory routes, by widening the dissected vessel area and including the vein in it.

    Have a nice summer!

    Best regards,

    Annamari

  • Rupa Lavanya Kaskurthy added an answer:
    Is there a free international pharmacological journal for publishing an article?
    Can you please reference a free journal regarding pharmacological, hormonal , antioxidant preclinical mammary cancer studies in which to publish? Ideally with a good impact factor???
    Rupa Lavanya Kaskurthy · BioPlus Life Science

    still i could not find the free one to publish my pre clinical herbal study...please kindly suggest

  • Fatih Yesildal added an answer:
    Is there a rabbit model for breast cancer by chemical carcinogenesis?

    We are planning a study about solid tumors and we need a rabbit model of chemically induced breast cancer but not cancer-cell-inoculated models.

    Fatih Yesildal · Gulhane Military Medical Academy

    Thanks for your answer. In our previous study, we had used NMU for breast cancer model in rats. The aim of that study was different. But in this study, we need frequent blood sampling, resulting in need for high amounts of blood. That's why we have planned a bigger animal for cancer model. Rats do not give us the opportunity of blood sampling in high amounts. If we used the same chemical in larger amounts (per kilogram) on rabbits, would it work? 

  • Prakash Dharmalingam added an answer:
    I am gonna inject MCF-7 cells to nude mices , do you have a suggestion for injection amount ? 1* 10^7 cells is enough or not ?
    and should I add Extra Fetal bovine serum, antibiotics or fetal bovine insulin during injection ?

    Hi-

    Using 10 million cells to inject in nude mouse for xenograft studies in practically Possibe. Is there any limitations in using that or any suggestions to be followed while using 10 milion cells for injection in nude mouse

     

    Thanking you

  • Richard Bright added an answer:
    Is there a way to induce metastaic breast cancer in normal mice (with thymus)?
    Generally metastatic breast cancers are induced in athymic mice using cell lines and maintaining the athymic mice is really challenging. I wish to know whether metastatic breast cancer can be induced in a normal mice (either through chemical inducer or cell lines or grafts). Please help.
    Richard Bright · University of Adelaide
    Orthotopic or ectotopic implantation of 4T1 cells in the skin or mammary fat pad, should form primary tumors followed by the formation of metastasis.
  • Gabriel Figueroa-Parra added an answer:
    How can you prevent cell clumping while preparing mouse splenocytes for adoptive transfer?
    I use cell strainer to mash the spleens and ACK lysis buffer or 9ml ddH2O (15sec) + 1ml PBS to lyse the RBC. But after spinning to collect cells, I would get cell clumps which are hard to break and that would cause me to lose the cells. Does anyone know a better method to prepare splenocytes where cell clumping is not a problem?
    Gabriel Figueroa-Parra · Autonomous University of Nuevo León
    We use this protocol, without any DNAse

    Good luck
  • Nirav M. Desai added an answer:
    Can I perform an ELISPOT assay on snap-frozen spleens?
    I snap-froze rat spleens and have stored them at -80C for the past month. Can I thaw the spleens and perform an ELISPOT assay after harvesting the splenocytes from the tissue? If so, how should I go about thawing and preparing the sample?
    Nirav M. Desai · Cadila Pharmaceuticals Ltd.
    I have worked with Frozen PBMCs and Spleenocytes with cryoprotectants. There is a loss in number though. Frozen spleen will not give you live cells to be used in ELISPOT.

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