Generation of antibodies directed against the low-immunogenic peptide-toxins microcystin-LR/RR and nodularin.
The preparation of antibodies against the liver toxin microcystin, as described here, is of major importance for its detection and purification in food and water, and for a therapeutic approach to neutralize the toxin by passive immunization. Microcystin-LR (MLR) and microcystin-RR (MRR) were purified from cyanobacterial cell materials by extraction, Sephadex LH-20-, ODS silica gel-, ionic exchange and RP-HPLC-chromatography. In order to reduce the toxicity for parenteral administration, microcystins were coupled by the carbodiimide method to poly-L-lysine (PLL(50.000)). Mice and rabbits were immunized with the conjugates in the presence of two lipopeptide immunoadjuvants (P(3)CSK(4) and P(3)CS-T(h)). High MLR-specific antibody levels were observed after parenteral coadministration of antigen and lipopeptides, whereas no anti-MLR antibodies were obtained with free microcystin or the microcystin-PLL(50.000)-conjugate in the absence of lipopeptide. In oral immunization, coadministration of antigen and adjuvants resulted in an accelerated development of anti MLR-specific antibodies and high antibody levels. Using the antisera, we could detect different microcystins and nodularin down to a concentration range of 10-50 ng/ml by a competitive inhibition ELISA; detection of microcystins in crude cell preparations was also possible. Furthermore, microcystins from different sources could be detected and discriminated from cyclic cyanopeptolines.
Baier
W
W
Institut für Immunbiologie der Universität, Stefan-Meier-Str. 8, D-79104, Freiburg, Germany.
Loleit
M
M
Fischer
B
B
Jung
G
G
Neumann
U
U
Weiss
M
M
Weckesser
J
J
Hoffmann
P
P
Bessler
W G
WG
Mittenbühler
K
K
D000284:Administration, Oral; D000818:Animals; D000917:Antibody Formation; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D007106:Immune Sera / Q000276:immunology; D007114:Immunization; D008387:Marine Toxins; D051379:Mice; D008807:Mice, Inbred BALB C; D052998:Microcystins; D010456:Peptides, Cyclic / Q000276:immunology* / Q000302:isolation & purification; D011817:Rabbits
D007106:Immune Sera; D008387:Marine Toxins; D052998:Microcystins; D010456:Peptides, Cyclic; C063998:nodularin; C078588:microcystin; C057862:cyanoginosin LR