ABSTRACT
Some unique subclasses of Camelidae antibodies are devoid of the light chain, and the antigen binding site is comprised exclusively of the variable domain of the heavy chain (VHH). The recombinant VHHs have a high potential as alternative reagents for the next generation of immunoassay. In particular, they might be very useful for molecular mimicry. The present study demonstrated an alpaca immunized with the F(ab')2 fragment of anti-aflatoxin B1 mAb and developed an important anti-idiotypic (anti-Id) responses. Antigen-specific elution method was used for panning private anti-Id VHHs from the constructed alpaca VHH library. The selected VHHs were expressed, renatured, purified, and then identified by a competitive enzyme-linked immunosorbent assay (ELISA). Our findings indicated that the VHH would be an alternative tool for haptens mimicry studies.
Subject(s)
Animals , Aflatoxin B1 , Allergy and Immunology , Amino Acid Sequence , Antibodies, Anti-Idiotypic , Chemistry , Camelids, New World , Allergy and Immunology , Immunoglobulin Heavy Chains , Chemistry , Molecular Sequence DataABSTRACT
<p><b>OBJECTIVE</b>To develop the methodology of gene chip to analyse genes involved in aflatoxin biosynthesis.</p><p><b>METHODS</b>In comparing reversed transcriptional PCR with gene chip, the gene chip was used to detect genes involved in aflatoxin biosynthesis.</p><p><b>RESULTS</b>After arrayed the slide was incubated in water for 2 hours, exposed to a 650 mJ/cm2 of ultraviolet irradiation in the strata-linker for 30 s, roasted under 80 degrees C for 2 hours in oven, pre-hybridized for 45 minutes and dealt with other procedures. Finally, the slide was hybridized with fluor-derivatized sample at 42 degrees C for 16 hours.</p><p><b>CONCLUSION</b>With the reasonable probe design and applicable protocol, the gene chip was prepared effectively for research on genes involved in aflatoxin biosynthesis.</p>