从老鼠血液通过大隐静脉和心脏穿刺“

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Summary

抽血是在大量研究的必要,例如,研究化合物的药代动力学轮廓。在这里,我们展示了如何绘制血液从大鼠使用两种技术:从大隐静脉或心脏穿刺抽血。

Cite this Article

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Beeton, C., Garcia, A., Chandy, K. G. Drawing Blood from Rats through the Saphenous Vein and by Cardiac Puncture. J. Vis. Exp. (7), e266, doi:10.3791/266 (2007).

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Abstract

绘制从啮齿类动物的血液,是必要的,无论在体外和体内研究的大量。网站绘制的血液是在啮齿类动物中数不胜数:眼窝窦,颈内静脉,上颌静脉,大隐静脉,心脏。每种技术都有其优点和缺点,和一些在一些国家没有批准任何更多的(例如,在荷兰的眼窝平)。讨论抽血不同的技术可用1-3。在这里,我们目前从老鼠的血液,两种技术各有其特定的应用程序。

从大隐静脉抽血,只要做得好,诱导动物最小的窘迫和不需要麻醉。这种技术可以反复吸引了少量的血,如需要4,5药代动力学研究,确定等离子体化学,或血球计数6。

心脏穿刺可收集大量的血液从单一的动物(最多10毫升血液,可以从一个150克的老鼠绘制)。因此,这种技术是非常有用的,作为一个终端程序,吸取血隐时将无法提供足够大的样本。我们用心脏穿刺,当我们需要从具体的增长在体外培养 4-9的T淋巴细胞的线条大鼠应变足够数量的血清。

Protocol

从老鼠血液通过大隐静脉和心脏穿刺“

:所有的程序必须由研究所的动物护理和使用委员会批准。

1。从大隐静脉抽血

  1. 执行此过程无需麻醉,需要两个人,一个人处理的大鼠,并执行平局。
  2. 走出一条毛巾,一个圆锥体或鼠与灭鼠处理手套,留下一个后肢暴露。
  3. 剃须电动微调的腿后面,直到大隐静脉可见。剃须一个足够大的面积,所以会在穿刺部位的接触,无毛。使用少量的非芳香洗手液,以保持非剃光头发远离穿刺部位。
  4. 压缩点的腿基地,使大隐静脉凸出(类似于使用止血带,从一个人或一个大型动物血液时)。使用20G针头静脉穿刺和血舀出来,因为它使用一个microvette。抽水的腿,将有助于吸引更多的血液。当你已经收集了足够的血液,保持一个干净的压缩穿刺部位止血。

2。心脏穿刺抽血

  1. 这个过程需要麻醉和终端。必须立即动物安乐死在心脏穿刺结束。
  2. 准备与23G1针5毫升注射器。
  3. 深anesthesize大鼠和麻醉检查缺乏自发的运动,呼吸速度缓慢,缺乏刺激(如捏脚趾)。为了确保长时间的麻醉,放置一个针筒,含有挥发性麻醉手术过程中浸泡在老鼠的鼻子的纸巾。
  4. 将大鼠在它的后面,面临着来自你走。
  5. 如果你是右撇子,你的左手食指放在水平最低的肋骨,不应用任何压力。将位于心脏〜1厘米,上面这点稍微向右。
  6. 保持在一个45度角的注射器,插入两根肋骨和手表之间的一滴血液来进针针。这是一个迹象表明,你在心里。而不移动您的注射器,拉填充注射器的柱塞。一旦注射器是全面的,仔细断开它从针头和空管。注射器就可以重新连接到针吸取更多的血液。它应该有可能借鉴从120-180克大鼠5-10毫升的血液。
  7. 立即安乐死的老鼠。

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Discussion

吸取大隐静脉的血液,是一种方便的方法得到少量的血,不需麻醉。如果需要重复抽样,检查法规,以确保您不画从一个老鼠太多血。

绘图血液由心脏穿刺是一种方便的方式来获得大量的血液,但是这是一个终端程序。被安乐死的动物必须在抽血结束。

这些技术(如活的动物所有的技术),应首先做一名兽医或兽医技术员,谁就能以评估动物是妥善处理,并不会受到不必要的痛苦和/或遇险的存在。

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Materials

Name Type Company Catalog Number Comments
Needle 20G 1/2 Tool BD Biosciences 305176 For puncturing the saphenous vein
Microvette 300 Tool Sarstedt Ltd 20.1308.100 To collect blood from the saphenous vein
Electric trimmer Tool Braintree Scientific, Inc. CLP-32130
Needle 23 G 1 Tool BD Biosciences 305145 For cardiac puncture
5 ml plastic syringe, slip tip Tool Fisher Scientific 14-826-12 For cardiac puncture

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References

  1. Van Herck, H., Baumans, V., Brandt, C. J. W. M., Boere, H. A. G., Hesp, A. P. M., Van Lith, H. A., Schurink, M., Beynen, A. C. Blood sampling from the retro-orbital plexus, the saphenous vein and the tail vein in rats: comparative effects on selected behavioural and blood variables. Laboratory Animals. 35, 131-139 (2001).
  2. Luzzi, M., Skoumbourdis, E., Baumans, V., Conte, A., Sherwin, C., Kerwin, A., Lang, T., Morton, D., Barley, J., Moreau, E., Weilenmann, R. F., Reinhardt, V. Collecting blood from rodents: a discussion by the laboratory animal refinement and enrichment forum. Animal Technology and Welfare. 4, 99-102 (2005).
  3. Angelow, O., Schroer, R. A., Heft, S., James, V. C., Noble, J. A comparison of two methods of bleeding rats: the venous plexus of the eye versus the vena sublingualis. Jounal of Applied Toxicology. 4, 258-260 (2006).
  4. Beeton, C., Wulff, H., Barbaria, J., Clot-Faybesse, O., Pennington, M., Bernard, D., Cahalan, M. D., Chandy, K. G., Beraud, E. Selective blockade of T lymphocyte K+ channels ameliorates experimental autoimmune encephalomyelitis, a model for multiple sclerosis. Proc. Natl. Acad. 13942-13947 (2001).
  5. Beeton, C., Pennington, M. W., Wulff, H., Singh, S., Nugent, D., Crossley, G., Khaytin, I., Chen, C. Y., Calabresi, P. A., Chandy, K. G. Targeting effector memory T cells with a selective peptide inhibitor of Kv1.3 channels for therapy of autoimmune diseases. Mol. Pharmacol. 1369-1381 (2005).
  6. Beeton, C., Wulff, H., Standifer, N. E., Azam, P., Mullen, K. M., Pennington, M. W., Kolski-Andreaco, A., Wei, E., Grino, A., Counts, D. R., Wang, P. H., LeeHealey, C. J., Andrews, B. S., Sankaranarayanan, A., Homerick, D., Roeck, W. W., Tehranzadeh, J., Stanhope, K. L., Zimin, P., Havel, P. J., Griffey, S., Knaus, H. G., Nepom, G. T., Gutman, G. A., Calabresi, P. A., Chandy, K. G. Kv1.3 channels are a therapeutic target for T cell mediated autoimmune diseases. Proc. Natl. Acad. Sci. USA. 17414-17419 (2006).
  7. Beeton, C., Barbaria, J., Devaux, J., Benoliel, A. -M., Gola, M., Sabatier, J. -M., Bernard, D., Crest, M., Beraud, E. Selective blocking of voltage-gated K+ channels treats experimental autoimmune encephalomyelitis and inhibits T-cell activation. J. Immunol. 936-944 (2001).
  8. Devaux, J., Forni, C., Beeton, C., Barbaria, J., Beraud, E., Gola, M., Crest, M. Myelin basic protein-reactive T cells induce conduction failure in vivo but not in vitro. Neuroreport. 317-320 (2003).
  9. Beeton, C., Chandy, K. G. Induction and monitoring of adoptive delayed type hypersensitivity in rats. Journal of Visualized Experiments. 8, (2007).

Comments

13 Comments

  1. DŒs the cardiac puncture procedure also work with mice? If not, how can I do a cardiac puncture with mice?

    Reply
    Posted by: Anonymous
    October 1, 2008 - 6:35 PM
  2. Yes, this cardiac puncture also works with mice. It is just a little more difficult as a ²0g mouse is about 10 times smaller than the rat shown on this video but, with a little practice, the mouse cardiac puncture can be done with consistent results.

    Reply
    Posted by: Christine B.
    October 1, 2008 - 6:42 PM
  3. Is it possible to draw 1 mL of whole blood from the Saphenous vein of a 150 g rat?

    Reply
    Posted by: Anonymous
    October 26, 2008 - 10:00 PM
  4. 1 ml is a lot of blood and I doubt you can get that much reliably from the saphenous vein. I usually draw 50-100 ul, sometimes up to 300 ml.

    Reply
    Posted by: Christine B.
    October 27, 2008 - 11:13 AM
  5. is there is any need to change the needle size for doing experiments in mice

    Reply
    Posted by: Anonymous
    November 10, 2008 - 11:08 PM
  6. I would try with a ²3G, like for rats. You can use ²5G but then you will start seeing more hemolysis.

    Reply
    Posted by: Christine B.
    November 11, 2008 - 4:47 PM
  7. Why isn't hemolysis acceptable?

    Reply
    Posted by: sumali k.
    January 4, 2010 - 2:24 PM
  8. Why isn't hemolysis acceptable?

    Reply
    Posted by: sumali k.
    January 4, 2010 - 2:25 PM
  9. Whether of not hemolysis during blood draw is acceptable depends on the use you will have for the blood.
    We often use blood drawn through terminal cardiac puncture for the preparation of serum for tissue culture. In this case we try to avoid hemolysis as this affects cell survival in culture. For other applications, hemolysis may not represent a problem.

    Reply
    Posted by: Anonymous
    January 4, 2010 - 2:34 PM
  10. How much heparin to you add to your tubes before collecting the blood?

    Reply
    Posted by: Alexis L.
    February 3, 2010 - 1:09 PM
  11. If you want to collect plasma, prepare a solution of 5% heparin in PBS and add 0.1 ml of it to each 3-5 ml syringe.

    Reply
    Posted by: Anonymous
    February 3, 2010 - 1:12 PM
  12. For vena saphena: If you puncture the vein lower (next to the ankle), it's easier, because the vein in this area is more on the surface.

    Reply
    Posted by: Anonymous
    August 17, 2011 - 3:45 AM
  13. I'm a graduate student from P.R.China.In our lab, we draw blood from mice from the retro-orbital sinus, which technique I find really disconcerting. I'm fascinated by your technique demonstrated here, and I would be grateful if Ms Beeton can send me the written protocol via e-mail.

    Reply
    Posted by: Anonymous
    January 31, 2012 - 7:32 PM

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