Evaluation of Newly Developed Dot-ELISA Test for Identification of Naja-naja sumantrana and Calloselasma rhodostoma Venom Antigens

Authors: A.S. Sikarwar, S. Ambu, T .H. Wong

Abstract:

Snake bite cases in Malaysia most often involve the species Naja-naja and Calloselasma rhodostoma. In keeping with the need for a rapid snake venom detection kit in a clinical setting, plate and dot-ELISA test for the venoms of Naja-naja sumatrana, Calloselasma rhodostoma and the cobra venom fraction V antigen was developed. Polyclonal antibodies were raised and further used to prepare the reagents for the dot-ELISA test kit which was tested in mice, rabbit and virtual human models. The newly developed dot- ELISA kit was able to detect a minimum venom concentration of 244ng/ml with cross reactivity of one antibody type. The dot-ELISA system was sensitive and specific for all three snake venom types in all tested animal models. The lowest minimum venom concentration detectable was in the rabbit model, 244ng/ml of the cobra venom fraction V antigen. The highest minimum venom concentration was in mice, 1953ng/ml against a multitude of venoms. The developed dot-ELISA system for the detection of three snake venom types was successful with a sensitivity of 95.8% and specificity of 97.9%.

Keywords: ELISA, Venom, SVDK, Naja-naja sumatrana , Calloselasma rhodostoma.

Digital Object Identifier (DOI): doi.org/10.5281/zenodo.1059785

References:

[1] Tan NH.Management of Snakebites in Malaysia
[Internet].
[cited 2010 Sep27].Availablefrom:http://www.tanngethong.com/Management%20of %20snakebites/management_of_snakebites_in_mala.htm.
[2] Alirol E, Sharma SK,Bawaskar HS,Kuch U,Chappuis F.Snake bite in South asia: a review.PLos Negl Trop Dis. 2012;4(1):e603.
[3] Kasturiratne A, Wickremasinghe AR, deSilva N,Gunawardena NK, Pathmeswaran A, et al.The global burden of snakebite:A literature analysis and modeling based on regional estimates of envenoming and deaths. PLoS Med 5. 2008;e218.doi:10.1371/journal.pmd.0050218
[4] World Health organization. Rabies and envenomings:A neglected public health issue.2007. Geneva:WHO.Available at http://www.who.int/blood products/animal_sera/rabies_envenomings/en/index.html
[5] Gutierrez JM, Theakston RDG, Warell DA. Confronting the neglected problem of snake bite envenoming: The need for a global partnership. PLoS Med.2003;3:e150.doi:10.1371/Journal.pmed.0030150.
[6] Warrell DA. Snake bite. Lancet. 2010 Jan 2;375(9708):77-88
[7] Engvall,E and P.perlman. Enzyme linked immunosorbent essay (ELISA): Quantitation of specific antibodies by enzyme labeled antiimmunoglobulin in antigen coated tubes. J.Immun.1971,109:129- 135.
[8] Heck FC,Williams JD, Pruett J. Interpretation of spectrophotometric absorbance values to define results of enzyme -linked immunosorbent assays. J Clin Microbiol.1980 Apr;11 (4):398-401
[9] Bechi P, Baldini F,Cianchi F. What is the reliability of the portal spectrophotometer with a fiberoptic probe to detect bilirubin in bile refluxate? Is sensitivity of Bilitec likely to be improved? Chapter, GER and gastric motility factors. Available at http://ww.hon.ch/OESO/books/Vol_5_Eso_Junction/Articles/art118.htm l