Use of affinity-purified antibodies to measure the in vivo disappearance of antibodies to myotoxin a

Abstract

M. A. Bober and C. L. Ownby. Use of affinity-purified antibodies to measure the in vivo disappearance of antibodies to myotoxin a. Toxicon 26, 301 – 308, 1988. — Antiserum against myotoxin a was purified using affinity chromatography. Myotoxin a was conjugated to an AffiGel agarose gel bead support and crude antiserum applied to the column. Antibody was eluted with distilled water, acetic acid and phosphate-buffered saline, and the protein concentration in the effluent was estimated by the absorbance at 280 nm. Antibody eluted with distilled water was used to develop an ELISA to detect antibodies to myotoxin in the bloodstream. Mice were injected with either 0.10, 0.15 or 0.20 ml of crude antiserum, and blood samples were taken during a four-week period. Samples were assayed for antimyotoxin using the antibody detection ELISA. Blood levels of antimyotoxin decreased significantly ( P < 0.05) within 1 hr after mice received 0.10 ml of crude antiserum (i.v.). Levels of antimyotoxin in mice given 0.15 and 0.20 ml of antiserum decreased significantly at 3 hr after the injection. Mice given 0.20 ml antiserum had significantly ( P < 0.05) higher amounts of antimyotoxin than mice receiving 0.10 ml antiserum during the 24-hr period after injection. However, after 24 hr all three treatment groups had less than 100 ng antimyotoxin/ml and did not differ significantly from one another. Measurement of antivenom in the bloodstream of snakebite patients might help determine if and when additional antivenom should be administered.