小鼠结肠炎模型使用葡聚糖硫酸钠(DSS)

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Summary

葡聚糖硫酸钠(DSS),在饮用水管理是一个既定的小鼠急性结肠炎的炎症损伤模型。该协议列出了DSS的治疗方法和组织编制。

Cite this Article

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Whittem, C. G., Williams, A. D., Williams, C. S. Murine Colitis Modeling using Dextran Sulfate Sodium (DSS). J. Vis. Exp. (35), e1652, doi:10.3791/1652 (2010).

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Abstract

结肠炎可发生病毒或细菌感染,缺血性侮辱,或自身免疫性疾病,最显着的溃疡性结肠炎和克罗恩病的结肠变种 - 克罗恩结肠炎。急性结肠炎患者可出现腹痛,腹胀,吸收不良,腹泻,便血和大便粘液。我们开始认识到环境,遗传学,和上皮屏障功能障碍在炎性肠病结肠炎动物模型已在推进我们对本病的认识至关重要之间复杂的相互作用。一种流行的模式涉及与低分子右旋糖酐硫酸钠(DSS)的补充对小鼠的饮用水,从而导致在结肠上皮损伤和一个强大的炎症反应,持续数天

Protocol

第1部分:损伤修复结肠炎模型

  1. 基线体重应获得每个鼠标之前开始直接资助的治疗。
  2. 设为3%(W / V)葡聚糖在水硫酸钠盐溶液,并用0.45μm的醋酸纤维素过滤器的过滤器。
  3. 四天的3%DSS解决方案,更换鼠笼的饮用水。这些老鼠不应该有任何其他来源的水(即排除放置在自动浇水系统的提示)。
  4. 第四天,一个额外的三天更换水DSS解决方案,允许一些结肠上皮复苏。 4天小鼠应权衡利弊,以量化结肠炎的系统性后果。减肥是常见的严重伤害。
  5. 第7天,权衡和牺牲的老鼠。小鼠可吸入过量的异氟醚,或其他制度上安乐死,IACUC批准的方法。

第2部分:剖检和科隆收获

  1. 揭露鼠标和安全的腿腹端,腹部,以确保通畅的访问。完全用70%乙醇湿腹部。
  2. 抓住组织钳腹部中线,从而延长向上露营的皮肤。使用细尖的剪刀切开腹部暴露腹腔内容。延长切口的xyphoid过程中提示在中线,然后沿伪劣方面的双边沿海的利润率延长。
  3. 确定小肠,盲肠和结肠。仔细解剖/挑逗从周围的肠系膜结肠。断面结肠深藏在骨盆免费的远端结肠和直肠。在colonocecal距断面免费近端结肠的结肠。必须小心,在此过程中,不损害结肠清洗结肠,将是有问题的的,如果发生穿孔。
  4. 使用20G喂养的针头和10 ml注射器,插管和冰冷PBS冲洗大肠,直到洗脱液是完全明确的大便。
  5. 在这一点上,如果想要的宏观分析,在解剖显微镜,冒号,可以固定为“香肠”。如果病理分析只需要进行第4部分“急性结肠炎组织学分析制作的瑞士卷”。

第3部分:处理整个结肠的宏观分析“香肠”

  1. 重要的是结肠保持正确的方向,因此,保持最接近运营商结肠远端。切两片非吸收缝合线(SP116 1.5公吨编织丝绸),一个约1英寸长,其他的2英寸。
  2. 使用2英寸片缝合,以配合末端接近尽可能的保证金,同时仍保持良好的密封。
  3. 放置一个20G的喂养针含5毫升10%福尔马林磷酸盐缓冲到近端的结肠。松散的领带喂养针灯泡立即近端缝合,其余部分。牢牢把握冒号在喂养针灯泡和注入福尔马林,直到结肠扩大。在缝合拧紧结,你退出喂养针,从而使作为“香肠”的注入,扩大结肠。
  4. 填写一个15毫升的锥形管,用10%福尔马林磷酸盐,并将其放置在管在结肠。修正为24小时。
  5. 倒入福尔马林和一个额外的24小时用70%乙醇代替。 (冒号可以存储在70%乙醇在室温下无限期。)
  6. 从锥形管中取出结肠和削减每年年底的字符串,用手术刀小心不要破坏结肠。请记住,末端的字符串最长的那块。削减纵向从远端到近端结肠,使之形成一个长表。此时,冒号可以被看作在解剖显微镜下。

第4部分:急性结肠炎的组织学分析的瑞士卷

  1. 重要的是结肠保持正确的方向,因此,保持最接近运营商结肠远端。
  2. 测量结肠长度。该指标损伤的严重程度的另一个指标。结肠炎增加水肿,并缩短整体结肠长度。
  3. 用细尖的剪刀,切割纵向从远端到近端结肠。用细尖镊子把握要么切口的边缘和横向打开远端,近端自己的方式工作,从而为平板显示结肠。
  4. 滚动结肠需要一个钳和一双双手技术。把握要么钳远端缘的边缘。顺序辊旋转并释放钳继续向近侧缘结肠。从而产生了第三个维度或“瑞士卷”的一个螺旋。要保持卷的形式,牢牢把握辊钳和断面与27G1 / 2针。安全针,弯针,因为它退出卷筒。
  5. 放置在一个适当的标记组织的录音带和一罐10%的轧辊,缓冲福尔马林磷酸盐在室温为24小时,以确保组织固定。
  6. 倒入福尔马林和一个额外的24小时用70%乙醇代替。 (冒号可以存储在70%乙醇在室温下无限期。)
  7. 一旦样品已被固定,石蜡包埋,切片和病理分析的幻灯片上。

第5部分:代表结果

急性结肠炎的DSS模型允许研究者获取,修复,和分析一个冒号,急性结肠炎模型。瑞士卷被切断,安装时,应该形成一个代表整个结肠切片,如果推出正确(图1)。安装成卷,可与H&E染色,以确定损害程度的结肠,从远端(内)结束的近端(外)结束(图2)。也可进行免疫组织化学瑞士卷,以确定和量化炎症浸润。如果香肠的方法已被正确地执行,将固定结肠扩张,整个粘膜表面,可以轻松地操纵下的解剖显微镜(图3)观察。

图1
图1,正确执行的“瑞士卷“。 (一)结肠推出从远端到近端,用针横断和弯针年底固定。它,然后放置在一个固定的组织卡带。 (二)H&E染色5微米的鼠标与决策支持系统(D =远端结肠P =近端结肠)治疗结肠作出了瑞士卷部分。

图2
图2:决策支持系统治疗冒号显示急性结肠炎迹象。炎症和隐窝损伤DSS治疗结肠水治疗控制比较明显。

图3
图3。“香肠”的一个例子。香肠用福尔马林注入完全展开。一个很小的角度,将目前中学的自然弧度的结肠。打开香肠应平放。

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Discussion

这个协议可以进行修改,以急性损伤,损伤修复,或慢性结肠损伤过程模型。

急性损伤的修改:

决策支持系统治疗4-6天,立即牺牲即兴

损伤修复的修改:

伤4-6天3-4天,和牺牲的恢复期间(如在上述协议中所述)的DSS的治疗。

慢性结肠炎的修改:

小鼠被放置在三个为期五天的3%DSS的周期,每个周期之间的复苏16天。小鼠牺牲后的最后16天的休息时间。

有几个问题,研究人员需要意识到这个模型:

  1. 在损伤部位的变异:在我们手中,我们看到一个远端为主的损伤模式,与近端结肠和盲肠的相对备用。其他有报道更近端的损伤模式的优势。
  2. 环境变化。有显着的环境变化。这是可能的,部分原因是由于肠道菌群,饮食和其他环境因素的差异。
  3. 这种模式是无效的,如果使用超高分子量DSS是(即50万大)。
  4. 是具有相当大的压力这种模式4变异。

生化分析结肠可抽取样本从近端或远端的利润。根据损伤的严重程度,这可能会影响你的能力产生病理损伤评分。在体内的BrdU标签来衡量扩散,可以通过腹腔注射16.7微克/公斤BrdU 2小时前牺牲α- BrdU免疫组化处理。

作为替代的“香肠”,冒号可以是固定的完全平坦的。线的Whatman纸与特百惠容器的底部,并浸泡于10%的纸张缓冲福尔马林磷酸盐。剖析在“第2部分:剖检和结肠癌的收获”中描述的结肠。再次,保持最接近运营商结肠远端。用细尖的剪刀,切割纵向从远端到近端结肠。随着细尖镊子把握或者边缘的切口和开放的横向合作,从而为平板显示冒号。转让本单位的工作表,预先浸泡的Whatman纸。盖上另一块的Whatman和湿夷为平地的结肠,用10%福尔马林磷酸。的滤纸应在福尔马林完全覆盖,但没有这么多,它升空结肠。密封的特百惠,24小时修复。从特百惠中取出结肠和10毫升70%乙醇15 ml锥形管。 (冒号可以存储在70%乙醇在室温下无限期。)

有几种方法量化结肠损伤5,6,7。一种方法,例如,使用一个多参数的规模,其中包括:炎症,参与程度,再生,隐窝破坏 ,以及第8%参与。

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Disclosures

范德比尔特IACUC规定的准则和法规的规定进行了动物实验。

Acknowledgements

由美国国立卫生研究院(1 K08 DK080221 - 01)和范德比尔特机构发展基金提供的资金。

Materials

Name Company Catalog Number Comments
Dextran Sulfate Sodium Salt USB Corp., Affymetrix 9011-18-1 mol wt 40,000-50,000
10% Buffered Formalin Phosphate Fisher Scientific SF100-4
Isoflurane, USP Phoenix Pharmaceuticals, Inc. NDC 57319-474-06
TISSUE PATH Macrosette Cassettes Fisher Scientific 15182706
BD 10 ml Syringe BD Biosciences 309604
Ethyl Alcohol Pharmco-AAPER E190 Dilute to 70% with distilled water
20G Straight Feeding Needle VWR international 20068-612
27G1/2 PrecisionGlide Needle BD Biosciences 305109
Dissecting Scissors Sharp/Blunt Tip VWR international 82027-588
Waugh Forceps VWR international 82027-428
Non-absorbable Suture LOOK Surgical SP116
Whatman Blotting Paper VWR international 28298-020 Cut as needed
Nalgene Surfactant-Free Cellulose Acetate (SFCA) Filter Cole-Parmer EW-06731-2
Carbon fiber composites digital caliper Fisher Scientific 15-007-958

DOWNLOAD MATERIALS LIST

References

  1. Okayasu, I. A novel method in the induction of reliable experimental acute and chronic ulcerative colitis in mice. Gastroenterology. 98, (3), 694-702 (1990).
  2. Martinez, J. A. Deletion of Mtgr1 sensitizes the colonic epithelium to dextran sodium sulfate-induced colitis. Gastroenterology. 131, (2), 579-588 (2006).
  3. Moolenbeek, C., Ruitenberg, E. J. The "Swiss roll": a simple technique for histological studies of the rodent intestine. Lab Anim. 15, (1), 57-59 (1981).
  4. Mahler, M. Differential susceptibility of inbred mouse strains to dextran sulfate sodium-induced colitis. Am J Physiol. 274, G544-551 (1998).
  5. Aharoni, R. Immunomodulatory therapeutic effect of glatiramer acetate on several murine models of inflammatory bowel disease. J Pharmacol Exp Ther. 318, (1), 68-78 (2006).
  6. Hahm, K. B. Loss of transforming growth factor beta signalling in the intestine contributes to tissue injury in inflammatory bowel disease. Gut. 49, (2), 190-198 (2001).
  7. Green, B. T. Adrenergic modulation of Escherichia coli O157:H7 adherence to the colonic mucosa. Am J Physiol Gastrointest Liver Physiol. 287, (6), G1238-1246 (2004).
  8. Dieleman, L. A. Chronic experimental colitis induced by dextran sulphate sodium (DSS) is characterized by Th1 and Th2 cytokines. Clin Exp Immunol. 114, (3), 385-391 (1998).

Comments

55 Comments

  1. what kind of mice you used in the experiment? hao ole of the mice when you treatment with DSS

    Reply
    Posted by: echo r.
    March 18, 2010 - 8:43 PM
  2. We use both 1²9 and C57BL6 mice in this experiment but the protocol can be used in any background. The mice are at least 6 weeks old before we begin treatment with DSS.

    Reply
    Posted by: Caitlyn W.
    March 19, 2010 - 10:27 AM
  3. what kind of mice you used in the experiment? hao ole of the mice when you treatment with DSS

    Reply
    Posted by: echo r.
    March 18, 2010 - 9:36 PM
  4. what happens if we keep the colon more than ²4 hours in 10%formalin ?

    Reply
    Posted by: peyman m.
    May 23, 2010 - 6:17 AM
  5. Leaving them in the 10% formalin shouldn't be a problem. We have left them over the weekend. For counts, longer fixation shouldn't be problematic, but I would put them in 70% ethanol before the end of 3 days.

    Reply
    Posted by: Caitlyn W.
    May 23, 2010 - 6:38 PM
  6. should DSS be prepared daily? and why it need be filtered?&#²9²59;Is the coulor of the solution&#²9²59;(DSS solute in filter water ) yellow

    Reply
    Posted by: Anonymous
    June 15, 2010 - 8:50 AM
  7. The DSS dŒs not need to be prepared daily. You can fill a water bottle enough for the four days on DSS. It needs to be filtered because you will be handling it before filtering and it can harbor bacteria, etc. that you don't want to feed to your mice. Yes, the color of the solution is slightly yellow.

    Reply
    Posted by: Caitlyn W.
    June 15, 2010 - 11:52 AM
  8. should I store the solution in routine temperature or in the fridge?

    Reply
    Posted by: Anonymous
    June 16, 2010 - 2:50 AM
  9. It is stored at room temperature.

    Reply
    Posted by: Caitlyn W.
    June 16, 2010 - 12:54 PM
  10. should I change the solution containing dss daily ? dŒs it influence the result if i do not change the solution daily?

    Reply
    Posted by: Anonymous
    June 16, 2010 - 9:48 PM
  11. There is no need to change the solution daily. Just put enough DSS solution in the water bottle on day one to last for the duration of the first cycle of DSS.

    Reply
    Posted by: Anonymous
    June 17, 2010 - 12:02 PM
  12. In our animal facility ,autoclaved tap water is used for all the mice. can I prepare DSS solution in tap water and then filter , or only distilled water shuld be used.

    Reply
    Posted by: Anonymous
    June 17, 2010 - 12:57 AM
  13. You can prepare the solution in tap water and then filter.

    Reply
    Posted by: Anonymous
    June 17, 2010 - 12:03 PM
  14. Dear Drs Whittem, Williams and Williams:

    Hello, I am writing a protocol to my University's IACUC to perform DSS induction of colitis, and was hoping that you may be willing to answer some of my questions regarding the animals subjected to DSS. I will be using the acute injury and injury repair models mentioned in your article. Could you please provide additional details of the stress that the mice might experience throughout the course of the experiment? The signs stated on the tutorial included weight loss, bloody stools, and diarrhea. Do these mice have additional issues such as an inability to retrieve food from an overhead feeding dish/water bottle, anorexia, inability to groom, contractures (i.e. 'hunching' in response to pain) or additional signs that may be useful to note when submitting my protocol? Also, should I expect mortality during treatment? As recommended, I will use mice older than six weeks, but likely not several weeks older as I want to avoid age-related issues. Finally, would you recommend the use of any type of analgesic drug during the course of DSS-colitis experiments to minimize discomfort in these animals? If so, would you happen to know the effect the analgesic has on factors such as barrier function, healing, infection rate, overall survival etc.? As I do not yet have any personal experience with this model, any insight will be greatly appreciated. Thank you.

    Reply
    Posted by: Elle F.
    June 28, 2010 - 8:03 AM
  15. For the acute injury and injury repair models you should only see the stresses mentioned in the article, weight loss, bloody stool, and diarrhea. There are no additional issues and you should not expect mortality during treatment. I would not recommend the use of an analgesic drug because, for example, NSAIDs will injure the gut and they should not be needed. Good luck!

    Reply
    Posted by: Caitlyn W.
    June 28, 2010 - 10:52 AM
  16. How do you utilize the sausages? I assume you slit them lengthwise after fixation and then stain (with what?) and examine them enface in a dissecting scope. What can you learn from this type of examination? Do you have a preferred method for quantitation of the resultand data?

    Reply
    Posted by: John W.
    July 30, 2010 - 3:05 PM
  17. You are correct, the sausages are cut lengthwise and stained with methylene blue and analyzed for aberrant crypt foci and polyp burden. The data is collected as and quantified as numbers of ACF/colon or number or size of polyps per colon.

    Reply
    Posted by: Caitlyn W.
    July 30, 2010 - 3:19 PM
  18. Read the abstract,
    "Mice are euthanized at the conclusion ... and at necropsy ... and can be 'Swiss rolled" 3 to allow microscopic analysis of the entire colon or infused with formalin as "sausages" to allow macroscopic analysis. Tissue is then embedded in paraffin, sectioned, and stained for histologic review."

    "Swiss roll" is for histological analysis, while "sausage" is for macroscopic analysis of the ENTIRE colon.

    Reply
    Posted by: Anonymous
    December 6, 2010 - 11:21 AM
  19. is dextran a reliable model for colitis...?do i use the drug for treatment before or after colitis using dextran?

    Reply
    Posted by: Anonymous
    June 12, 2011 - 7:20 AM
  20. DSS is one of the better models for colitis. The papers found at the following sites as well as those that we cite in our paper give good information on the topic. http://www.ncbi.nlm.nih.gov/pubmed/9844047?dopt=Citation http://www.ncbi.nlm.nih.gov/pubmed/8881816?dopt=Citation http://www.ncbi.nlm.nih.gov/pubmed/14611673?dopt=Citation We have not performed treatments on mice on DSS. We mainly use it to analyze the response of knockout models to experimental colitis. Treatment times would depend on whether you would like to prevent or treat colitis.

    Reply
    Posted by: Caitlyn W.
    June 15, 2011 - 6:24 PM
  21. I'm working with this model, but using ²% DSS as described. We bought our chemicals from another company, but the molecular weight is the same. Can you tell me more about the bloody diarrhea. I have finished the DSS water, but did not notice any blood. There was a bit of an excessive amount of feces, but I did not see the mice hair (white) or bedding stained with dried blood. I've read in other papers that not all the mice have it, but dŒs the fact that I did not see any mean that something went wrong?

    Reply
    Posted by: Anonymous
    July 17, 2011 - 1:21 PM
  22. There can be considerable variability in the severity of colitis induced by DSS. This depends on manufacturer, lot #, environment, differences in enteric flora. We typically track weights, DAI which includes stool consistency, blood, etc, measure colon length and weight at necropsy and for us, we consider the histology observed from reviewing the H+E's as the gold standard for injury. I would recommend reviewing the histology with a GI pathologist.

    Reply
    Posted by: Anonymous
    July 20, 2011 - 3:00 PM
  23. I am submitting a protocol for DSS to the IACUC. They want to know how I "will deal with the diarrhea with occult blood". Is there anything that we can administer to the mice to make the IACUC happy? They want to know how long I will let these symptoms go on before removing the mouse from the study. Unfortunately, all of these things are facets of the disease.

    Reply
    Posted by: Anonymous
    August 18, 2011 - 10:09 AM
  24. Hello Jesse,
    We insure that IACUC knows that mice that drop below ²0% weight loss are sac'd. The amount of time that mice are on DSS results in diarrhea and blood for only 4-6 days and mice recover well. You should not have to worry about mice having symptoms for more than a week.

    Reply
    Posted by: Caitlyn W.
    August 18, 2011 - 12:00 PM
  25. Thank you for your response. Another question...if the mice are in distress(eg. bloody diarrhea) but we don't give them any analgesic, then the IACUC will make them go down as Category E by USDA standards. DŒs anyone else have experience with this? DŒs anyone else list them as Category E?

    Reply
    Posted by: Anonymous
    August 18, 2011 - 2:00 PM
  26. Ours are Category E

    Reply
    Posted by: Caitlyn W.
    August 18, 2011 - 6:19 PM
  27. The water to be used with the DSS should be autoclaved water?

    Reply
    Posted by: Lilian N.
    August 18, 2011 - 10:50 PM
  28. DŒs anybody do anything to combat dehydration in these mice?

    Reply
    Posted by: Anonymous
    August 19, 2011 - 10:56 AM
  29. Sucrose 6%.
    http://farncombe.mcmaster.ca/documents/Ghiaetal.GastrŒnterology²0091367²²80-²²88.pdf

    Reply
    Posted by: Lilian N.
    August 19, 2011 - 12:57 PM
  30. The water used with DSS dŒs not need to be autoclaved. You should filter it after making the DSS solution. We do IP saline injections.

    Reply
    Posted by: Caitlyn W.
    August 19, 2011 - 1:39 PM
  31. DSS will be degrade or not in water. How DSS solution will be filtered what is procedure for that.can u elaborate thanks

    Reply
    Posted by: Anonymous
    September 24, 2011 - 3:11 AM
  32. We have not officially tested whether DSS degrades in water over extended periods of time. We make fresh DSS solutions for each cycle of DSS that is administered and do not reuse the DSS that is remaining in the water bottles after each cycle. As mentioned in the protocol, we make a 3% (w/v) Dextran Sulfate Sodium Salt solution in water and filter with a 0.45 μm cellulose acetate filter (Nalgene Surfactant-Free Cellulose Acetate (SFCA) Filter, Cole-Parmer, catalog number: EW-06731-²). The directions for this filter come with it but it is simply a vacuum filtration system as pictured in the video (seen at 0:5², "Injury-Repair Colitis Model").

    Reply
    Posted by: Caitlyn W.
    September 25, 2011 - 3:50 PM
  33. Has anyone used buprenorphine in the DSS model?

    Reply
    Posted by: Anonymous
    September 27, 2011 - 1:29 PM
  34. Question: In this mouse model of DSS colitis, do we have to to remove the colon contents (stools) before histological analysis? How is this done? Thank you!

    Reply
    Posted by: Anonymous
    October 14, 2011 - 10:49 AM
  35. At 3:17 in the video the removal of stools is shown. The colon is flushed with PBS using a ²0G straight feeding needle.

    Reply
    Posted by: Caitlyn W.
    October 17, 2011 - 4:38 PM
  36. In some protocols, people sacrifice the mice 3 or 5 days after DSS treatment. What is the purpose of this? How is this different from killing the mice after 4 days of DSS treatment.

    Reply
    Posted by: Anonymous
    January 16, 2012 - 4:52 PM
  37. The number of days that you sac after DSS treatment will determine the amount of time the mice have to heal. we generally use 4 days because, in wild type mice, this is usually the best amount of time to see the beginnings of recovery while still noting damage in the mice of interest used in our lab. Basically, this is the point at which we see the most difference between the two groups. If you have a mouse that you expect to heal more quickly, 3 days may be suitable, and vice versa.

    Reply
    Posted by: Caitlyn W.
    January 18, 2012 - 1:21 PM
  38. Regarding the step 7 of part 4 in the protocol mentioned above, how do you remove a needle from a swiss-rolled tissue before going forward to paraffin-embedding? Just pull out? Do you have a specific way to do that?

    Reply
    Posted by: Anonymous
    January 17, 2012 - 6:05 PM
  39. Once the tissue is formalin fixed, the tissue will be stiff enough that you can either unbend or clip the needle and slide it out of the roll without altering the shape of the Swiss roll. Our tissue processing core knows how to handle our tissue, but if you need to do it yourself, insure that the tissue has been in 70% ethanol at least overnight and then use gloves and forceps to remove the needle from the roll.

    Reply
    Posted by: Caitlyn W.
    January 18, 2012 - 1:24 PM
  40. What are the strains of mice used for this model? And what is the health status of the animals?

    Reply
    Posted by: Anonymous
    February 10, 2012 - 11:16 AM
  41. We use C57Bl6 mice for our experiments. Our mice are healthy before treatment with DSS.

    Reply
    Posted by: Caitlyn W.
    December 13, 2012 - 1:04 PM
  42. Hello, I am writing a protocol to my University's IACUC to perform DSS induction of colitis.Should I put DSS in Hazardous material form?Is there any method to detect the DSS excretion in feces and urine?How can we deal with the waste from DSS (unused solution)?

    Thanks,

    Reply
    Posted by: Jinggang L.
    December 13, 2012 - 11:22 AM
  43. Our IACUC protocol dŒs not include DSS as a hazardous material and we pour waste DSS down the drain. As far as I know, there is not a method to detect DSS excretion in feces and urine.

    Reply
    Posted by: Caitlyn W.
    December 13, 2012 - 1:06 PM
  44. Hi
    In the materials you list Dextran Sulfate Sodium Salt USB Corp., Affymetrix 9011-18-1 mol wt 40,000-50,000.
    However when I check Affymetrix website, this catalog number belongs to a MW: 500,000 product (http://www.affymetrix.com/esearch/search.jsp?pd=131111&N=4²94967²93) . is this what you used or otherwise what is the correct catalog number. thanks!

    Reply
    Posted by: selcen o.
    May 6, 2013 - 10:50 AM
  45. I am so sorry for the confusion. The number listed in the protocol is the CAS# which is similar between the 500,000 and 50,000 M.W. DSS. The actual catalog number is 14489. This number should get you what you need (http://www.affymetrix.com/estore/browse/brand/usb/product.jsp?productId=130²61#1_1).

    Reply
    Posted by: Caitlyn W.
    May 6, 2013 - 12:00 PM
  46. Thanks a lot for the catalog number!
    Could you please explain the injection of saline to prevent dehydration? How often and how much should it be injected? I guess all mice should get the same amount at the same time, right?
    And when the mice get sick, should we put some food on the bottom of the cage, or are they usually capable of getting the food themselves? thanks again.

    Reply
    Posted by: selcen o.
    May 13, 2013 - 3:44 PM
  47. We usually only inject mice that are not looking healthy and we generally do this as needed. I will only inject a 1ml bolus on a single day. You can repeat this the next day, but this is really just a last resort. We sacrifice them once they reach ²0% weight loss so they don't have undue stress. When the mice get sick, I do tend to try and put food in the bottom of the cage in case the mice aren't able to reach up into the food distributor. If the mice are getting so sick that you need to inject with PBS or move the food down, you may want to try a smaller concentration of DSS (²% or 1%) or perform the treatment for a shorter time. We have had several genotypes that don't handle the DSS as well and decreasing the concentration or treatment time really helps them. There is also facility to facility variability dependent on mouse microbiomes so my mice may be less sensitive than your mice to DSS and even your WTs might have different responses than mine. In short, we often need to tweak the protocol so that we can get injury while insuring survival for the extent of the protocol that we need to perform.

    Reply
    Posted by: Caitlyn W.
    May 13, 2013 - 4:29 PM
  48. thanks a lot, Caityln, you are really helping a lot because we have no experience in that.

    Reply
    Posted by: selcen o.
    May 14, 2013 - 4:56 AM
  49. Inducing colorectal cancer in rats: I am in a predicament; instead of the DSS Mw 40,000, I have ordered DSS (Mr 5,000) by accident. Unfortunately I noticed this too late and have a very limited time. I have read that the Mw of DSS does play a role in inducing colitis, whereby 5 kDa could induce milder form of colitis. I plan to inject my rats with AOM (12 mg/Kg), expose the rats to 2 cycles of 1 week of 2% DSS following 2 weeks of water. These rats are quite old, 12 weeks to be exact. They are also Wistar rats so at this time they are prone to develop cancer easily. However, I would like to know is using 5 kDa DSS worth it? Could you please provide me with some advice?

    Reply
    Posted by: Lynn C.
    February 16, 2014 - 6:30 AM
  50. Unfortunately, I do not think that Mr 5,000 will work.

    Reply
    Posted by: Christopher W.
    February 19, 2014 - 9:21 AM
  51. can we use dss with m.w 500.000 for induce colitis?

    Reply
    Posted by: flora s.
    January 29, 2015 - 9:41 PM
  52. Is there a difference between female and male mice? Which would should i choose?

    Reply
    Posted by: stu a.
    February 17, 2015 - 11:42 AM
  53. We have not noticed a difference with gender; however, as the environment is a strong contributor to the severity of colitis, I would recommend test experiments with male and female WT mice in your facility.

    Reply
    Posted by: Christopher W.
    February 17, 2015 - 12:24 PM
  54. Would you also have an advice on how to minimize the variability between the cages? I was considering to mix treated and not treated animals in the same cage (our treatment is injected). However I am afraid that the groups will have cross effects on each other.
    Thank you

    Reply
    Posted by: stu a.
    February 17, 2015 - 2:25 PM
  55. Is it plausible to use 500,000 mw DSS? 1-2% for induction oftumours with AOM, I thank all advice.

    Reply
    Posted by: VERONICA M.
    May 14, 2015 - 5:04 PM

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