If you just want to understand hemolysis assays then this paper from the Canadian blood service is quite good comparison of measurement methods http://www.ncbi.nlm.nih.gov/pubmed/22750731 . But to fully understand hemolysis activity between species of snake venoms it's beneficial to perform enzyme assays for Phospholipase and proteases do understand the differing components between species

1. Need this article

HTTP://WWW.SCIENCEDIRECT.COM/SCIENCE/ARTICLE/PII/S0041010101001039

Novel in vitro assays for assessing the haemorrhagic activity of snake venoms and for demonstration of venom metalloproteinase inhibitors

A Beea, , R.D.G Theakstonb, R.A Harrisonb, S.D Cartera

39 (9),2001: 1429–1434

If you just want to understand hemolysis assays then this paper from the Canadian blood service is quite good comparison of measurement methods http://www.ncbi.nlm.nih.gov/pubmed/22750731 . But to fully understand hemolysis activity between species of snake venoms it's beneficial to perform enzyme assays for Phospholipase and proteases do understand the differing components between species

Ida S. Sano Martins,

I would like to know the snake species you are interesting. Depending on the species there is an article more adequate.

I may suggest to use the hens'egg assay i.e.: Sells PG, Richards AM, Laing GD, Theakston RD. The use of hens' eggs as an alternative to the conventional in vivo rodent assay for antidotes to haemorrhagic venoms. Toxicon 1997;35(9):1413-21.

It worked very well, in fact we utilized for our last study on crotalus atrox liophylized reconstituted venom ( Calzia D, et al., J Biochem Mol Toxicol. 2011 Nov-Dec;25(6):377-81. doi: 10.1002/jbt.20404.),

Dear All, thanks a lot for help,

I am working on snake venoms of Naja Naja, Krait, Russel Viper

Can you please send me those articles if possible because my institute doesn't have subscritoptn for these journals

I am currently working on In-vitro assay of snake venom

see references:

1) Thromb Haemost. 1997 Nov;78(5):1372-80.

2) Biochem Pharmacol. 2002 May 1;63(9):1589-97.

3) Toxicon. 2007 Jul;50(1):153-65.

4) Toxicon. 2007 Sep 1;50(3):400-10.

5) J Venom Res. 2011;2:11-6.

6) Toxicon. 2011 Mar 15;57(4):580-5.

7) Biomed Pharmacother. 2010 Oct 23.

if you still have any doubts, feel free to contact me.

@andre Fuly

Dear Sir, Thanks a lot, some of the articles are paid, can you email me if possible on this id [email protected]

Thank you,

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3306279/

Dear Friends Need this article

Toxicon. 2002 Jun;40(6):659-65.

Snake venom L-amino acid oxidases.

Du XY, Clemetson KJ.

Source

Theodor Kocher Institute, University of Berne, Freiestrasse 1, CH-3012 Berne, Switzerland.

Erratum in

Toxicon 2002 Sep;40(9):1381.

Dear, you can find all these in Vivo, as well as in Vitro official techniques in WHO publication submitted by Dr. R.D.G. Theakston et. al

Dear Win,

I found this article from your suggestion are you talking about this one only?

Thank you,

Erratum to Report of WHO workshop on the standardization and

control on antivenoms” [Toxicon 41 (5) (2003) 541–557]q

R.D.G. Theakston*, D.A. Warrell, E. Griffiths

I need specific articles which can be used to know the hemolytic property of Snake venom............

hi, if it is too difficult to find on ur topic of interest (i.e. in vitro aasay of snake venom using nanoparticles), u can find so many articles related to in vitro assay of snake venom using plant extract. u can use these articles as references n develop protocol.

plz check following article:

Diversity of Micrurus Snake Species Related to Their

Venom Toxic Effects and the Prospective of Antivenom

Neutralization

Gabriela D. Tanaka1, Maria de Fa´tima D. Furtado2, Fernanda C. V. Portaro1, Osvaldo Augusto

Sant’Anna1, Denise V. Tambourgi1*

Hypericum brasiliense plant extract neutralizes some biological effects of Bothrops jararaca snake venom

Mariane Assafim, Eduardo Coriolano de Coriolano, Sérgio Eufrázio Benedito, Caio Pinho Fernandes, Jonathas Felipe Revoredo Lobo, Eladio Florez Sanchez, Leandro Machado Rocha, and André Lopes Fuly*

Snake venom neutralization by Indian medicinal plants (Vitex negundo and Emblica officinalis) root extracts

M.I Alam

all the best!

many enzymes and nonemzymatic components of snake venoms can cause red blood cells hemolysis ex:phospholipases nucleotidases cytolytic components etc..

I recommend going through internet searching for the components found in your cobra venom. then decide what you really want. It is just total hemolysis which may differ from snake to the other depending on what component is present.. hemolysis measurment is just very simple tech depends on destruction of washed blood cells and refer persenting hemolysis to a sample 100% hemolysed by water

Al-Abdulla, I.H.; Sidki, A.M. and Landon J. (1991): An indirect haemolytic assay for assessing antivenoms. Toxicon, 29: 1043-1046.

i use this assay to calculate specific hemolytic activity of venoms . the assay was peformed as follow:

100 µl of venoms were incubated with a pre-warmed assay mixture (37 °C) containing 100 µl of egg yolk suspension (diluted 1:5 in 0.9% isotonic saline), 1 ml of washed human erythrocytes [2.5% (V/V)] and 3.8 ml of hemolytic assay buffer (1 mM Tris-HCl buffer, pH 7.5 containing 100 mM NaCl, 100 mM KCl and 10 mM CaCl2) at 37 °C for 30 min. Venom was excluded from the blank tube. In order to have a 100% haemolysis value (control), erythrocytes were incubated with distilled water.

After incubation, the tubes were centrifuged at 4000 rpm for 15 min and the degree of haemolysis was determined by measuring the absorbency of the supernatant at 540 nm. The % haemolysis was calculated from the following equation:

% Haemolysis = (OD540 Venom/OD540 Control) × 100.

The amount of venom that gives 80% haemolysis was considered as one hemolytic unit, and the specific hemolytic activity (H.A.) of venoms was expressed as units/mg.

i hope i help u .

Thanks a lot Willo sir

and thanks to all

I think this article will be helpful for you to study the in vitro activity of the compound against snake venom

First you have to be clear for what type of snake venom you going study. Because you know that each differ in their activity. in case if you plan to do invitro PLA2 activity means you should be very clear in venom type because PLA2 vary from viper and cobra family. Viper family contain external c-terminal region in their structure which shown pharmacological site of the enzyme but in cobra and krait which have N-terminal region. These difference in their pharmacological sites shown in most of the enzymes nd proteins in venom.

anti-hemolytsis, palates studies and PLA2 inhibition studies are most important for invitro antivenom study. apart from this you have to be clear study about ur drug activity in particular tissue interaction of the venom, In simple you can use Student organ bath method to study all these activity. For cobra family venom mussels cell interaction study is very important . I have collected nearly 700 papers about snake venom studies. you can read one book " Hand book of venom and toxin of reptiles" there you can clearly understand the mechanism of differ enzymes in snake venom. If you give me ur mail id then i will forward those books. All the best for your research work.

I suggest you two simple and reliable methods. 1) the phenol red and 2) the indirect hemolytic test using egg yen´s yolk. If you want to try them, fell free to contact me.

Animal venoms can induce hemolysis by different ways. One is direct route, as in cases of Elapidae snake venoms and bees. In these cases there is a component in the venom which induces hemolysis directly affecting membrane of erythrocytes. Another pathway is the indirect hemolysis induced mainly by phospholipases present in Viperidae snake venoms. In this case, for testing in vitro you need to add lecithin to get lysolecithin that will act on the membrane. There are some works that explains these mechanisms. In addition, there is another indirect mechanism that you can find in some spiders venom that induces hemolysis via C3 complement. Then, it is important to know what you are thinking to do in order to help you …or not. best wishes, Ida

@sakthi ..........Can you please email me

my id is [email protected]

Thank you,