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An enzyme immunoassay for usnic acid in lichens was developed, the sensitivity of which was 0.1 microg/g of air-dried material (0.00001%). Polyclonal rabbit antibodies against bovine serum albumin conjugated to (+)-usnic acid under the conditions of formaldehyde condensation made it possible to determine the analyzed substance in solutions at concentrations from 1 ng/mL when it interacts with an immobilized gelatin conjugate homologous in the binding mode. Usnic acid in 2-26600 microg/g (0.0002-2.6%) amounts was found in all 236 studied samples of lichens belonging to 53 species and 8 families.

The composition of low-molecular biologically active metabolites typical of microscopic fungi has been studied in blastemas of fruticose lichens of the genera Cladonia, Cetraria, Evernia, Bryoria, and Usnes. The enzyme immunoassay method showed the presence of sterigmatocystin, emodin, mycophenolic acid, citrinin, alternariol, and diacetoxyscirpenol, which occurred regularly and, in most cases, at a frequency of 55 to 100%. The highest levels of accumulation were 0.001-0.003% for emodin, 0.0002% for alternariol and citrinin, 0.0001% for sterigmatocystin and mycophenolic acid, and 0.00005% of the weight of air-dry material for diacetoxyscirpenol. Other metabolites (cyclopiazonic acid, ergot alkaloids, ochratoxin A, PR toxin, deoxynivalenol, zearalenone, and fumonisins) were detected in these lichens less frequently (sometimes only upon the expansion of the territory of sampling), and their content was no more than 0.00005%. The peculiarities of the component composition and the levels of accumulation of fungal metabolites in lichens of different taxonomic affiliation were discussed.

A highly specific test system based on polyclonal rabbit antibodies targeting a conjugate of alternariol and bovine serum albumin prepared by formaldehyde condensation has been developed for the first time and shown to have the sensitivity of 0.4 ng/ml in the indirect competitive determination of alternariol. The possibility of applying the assay for assessment of the intensity of alternariol contamination of corn, animal feed, and raw materials used for the preparation of the latter has been demonstrated.

Using the reaction of activated N-hydrooxisuccinimide ester of mycophenolic acid, a series of immunoreactive conjugated antigens with albumins, gelatin, and glucosoxidase is obtained. On the basis of polyclonal rabbit antibodies, a test-system for indirect competitive immunoenzyme analysis is elaborated, which has the sensitivity 0.4 ng/ml. By immunoanalysis, the ability for active biosynthesis of mycophenolic acid in strains of Byssochlamys nivea (44/44, 4100-68400 ng/ml) and Penicillium roqueforti (7/16, 204-25120 ng/ml) from the mycobiota of ensiled feeds is confirmed. The correspondence between weakly expressed producing capacity of most species of fungi of the genera Penicillium and Aspergillus prevailing in grain feeds and the data on low occurrence of this metabolite in grain (8.0%) and combined feeds (11.9%) is confirmed. A potential relationship between particular cases of a significant accumulation of mycophenolic acid (from 500 to 1500 microg/kg) in grains of wheat, corn, and combined feeds and a high biosynthetic activity in rare species P. puberulum, P. stoloniferum, and P. gladioli is discussed.

The use of an indirect competitive enzyme-linked immune sorbent assay (ELISA) involving polyclonal rabbit antibodies against BSA-conjugated PR-toxin (sensitivity, 1 ng/ml) established the ability to synthesize PR-toxin in 18 out of 35 morphologically identified strains of Penicillium roqueforti and P. chrysogenum. The results indicate that ELISA for PR-toxin may be used in assessing the taxonomical position of terverticillate penicillia in the presence of other micotoxins.

Products of spontaneous conjugation of aflatoxins B1, G1, and G2 with bovine serum albumin (BSA) were shown to interact with antibodies against aflatoxins. Solid-phase BSA conjugates inhibited the binding of aflatoxins by anti-aflatoxin antibodies. Antisera against BSA-B1, BSA-G1, and BSA-G2 were obtained and their specificity, determined. The mechanisms of spontaneous binding of aflatoxins by proteins are discussed.

Aflatoxin B2a (AB2a), aflatoxin G2a (AG2a), and hemiacetal of sterigmatocystin have been shown to form immunoreactive conjugates with albumin. The conjugates were formed following incubation of solution mixtures at room temperature for 1 h, as demonstrated by spectrophotometry and enzyme immunoassay. Anti-AB2a antibodies reacted with AB2a, aflatoxin B1, and aflatoxin AB2 (100, 8.8, and 5.9%, respectively); a similar result was obtained for anti-AG2a antibodies reacting with AG2a, aflatoxin G1, and aflatoxin AG@2 (100, 2.5, and1.0%, respectively). Binding of anti-AB2a and anti-AG2a antibodies to solid-phase conjugates of AB2a or AG2a exhibited similar analytical characteristics.

Aflatoxin B1 and sterigmatocystine hemiacetal derivatives were synthesized, and their conjugation to albumins and gelatin and also spectral and immunochemthe specificity and analytical properties of the antibodies produced by immunization with conjugated antigens. The possible mechanism of hemiacetal interaction with proteins is discussed. Based on immune reagents to sterigmatocystine hemiacetal, a test system was developed for determination of sterigmatocystine at the sensitivity of 0.1 ng/ml.

The time course of production, specificity, and analytical potential of anti-zearalenone polyclonal rabbit antibody synthesized by formaldehyde condensation and conjugated to bovine serum albumin were investigated. The relative cross-reactivities with natural analogues were: zearalenone, 100%; alpha-zearalenol, 0.15%; and beta-zearalenol,0.02%. With synthetic analogues: zearalanone, 31.7% and alpha-zearalanol, 0.12%. An enzyme immunoassay for zearalenone with a sensitivity of 0.1 ng/ml was developed on the basis of these antibodies and solid-phase conjugates homologous to the immunogen in the method of synthesis.

Formation kinetics, specificity, and analytical potential of polyclonal antibodies raised in rabbits against BSA conjugates of carboxymethyloxime-zearalanone (CMO-ZAN) and carboxymethyloxime-zearalenone (CMO-ZEN). Preparation of the conjugates involved conversion of CMO-ZAN and CMO-ZEN into activated esters and carbodiimide condensation. Two versions of a group-specific enzyme immunoassay (for zearalenone/alpha-zearalenone and zearalanol/alpha-zearalanol) based on heterologous combination of solid-phase antigens are described (sensitivity, 0.01 ng/ml).

Four hybrid clones (MM-(AB1)-1, MM-(AB1)-2, MM-(AB1)-3, and MM-(AB1)-4) were obtained by hybridoma technology with immunization of BALB/c mice with a BSA conjugate of aflatoxin B1 carboxymethyloxime derivative. Antibodies produced by these clones varied in the ability to recognize the aflatoxin B1 analogues. The sensitivity of enzyme immunoassay based on all monoclonal antibodies was higher compared to analysis based on polyclonal rabbit antibodies (0.1 and 0.4 ng/ml, respectively).

Polyclonal rabbit antibodies against a conjugate synthesized through condensing BSA and disubstituted roridin A hemisuccinate allowed roridin A to be determined in solutions at a sensitivity of 0.2 ng/ml. The cross-reactivity of structural analogues--roridin A, verrucarin, and verrucarol--amounted to 100, 2.5, and 0.03%, respectively. The data showed that these antibodies determine roridin A in an indirect heterogeneous enzyme immunoassay in cereal straw samples at a sensitivity of 20 micrograms/kg.

Ochratoxin A was quantitatively monitored in grain extracts by indirect solid-phase enzyme immunoassay with the use of an immobilized conjugate of the toxin with gelatin and polyclonal rabbit antibodies raised against the ochratoxin A-BSA conjugate. This monitoring found that 1.7 to 18.5% of the samples were contaminated with the toxin at a concentration of 25.9-291.7 micrograms/kg. An analysis of forage grain found ochratoxin A at concentrations of 440-3250 micrograms/kg.

Indirect enzyme immunoassay based on immobilized conjugate of aflatoxin B1 carboxymethyloxime with bovine serum albumin and polyclonal rabbit antibodies allows determining aflatoxin B1 with a low relative cross-reactivity against aflatoxin B2, G1, G2, M1, B2a and G2a and sterigmatocystin (15.5, 15.5, 1.7, 1.0, 0.03, 0.03 and 0.01%, respectively) with a sensitivity of 0.04 ng per well or 4.0 ng per ml organic solvent.

Based on indirect solid-phase competitive enzyme immunoassay, a method for determination of T-2 toxin in grain was designed. Determination errors were measured on samples of contaminated grain. The method makes allows determination of the toxin levels ranging from 30 to 1000 ng/g.

A varriant of competitive ELISA, making it possible to detect fumonisin B1 (up to 0.4 ng/well) was developed, based on the use of specific polyclonal antibodies obtained by immunization of rabbits with a horseradish peroxidase conjugate of fumonisin B1. The minimum concentration of fumonisin B in water-acetonitrile extracts of corn grain, dry corn products, and feeds, detected by the ELISA version developed is 0.2 mg/kg.