Low cost production and purification of polyclonal antibodies to staphylococcal enterotoxin A

ABSTRACTAn immunization scheme for production of antiserum to staphylococcal enterotoxinA (SEA) is proposed. The reference method of Robbins and Bergdoll was modified toreduce the number of doses and the amount of toxin used per animal. The bestimmunization scheme used injections in days 0, 8, 24, 59, 62 and 67. The amount oftoxin at each injection was 5, 6, 20, 50, 100 and 200 µg, respectively. The total amountof toxin was 381µg, which corresponded to a reduction of 107µg in the amount oftoxin for each animal when compared to the reference method. The average antiserumtiter using the Optimum Sensitivity Plate - OSP was 1:60 and using ELISA the titerwas 1:100.000. The lack of cross-reactivity with other staphylococcal enterotoxinsindicated high specificity of the antibody to SEA. The proposed immunization schemewas adequate to produce specific SEA antisera, with high titers and the aditionaladvantage of reducing the amount of purified SEA required for immunization.Key words: Staphylococcus aureus, enterotoxins, detection, immunizationINTRODUCTIONStaphylococcal food poisoning is a worldwideintoxication caused by the ingestion ofstaphylococcal enterotoxins (SEs), preformed in foodby some Staphylococcus aureus strains. AlthoughS. aureus can be easily detected in foods, neither itspresence necessarily indicates enterotoxinproduction nor the absence of viable staphylococciassures food safety (4).The organism looses viability rapidly after thestationary phase, being replaced by harmlesssaprophytic bacteria. However, the toxins resistboth heat treatment and proteolytic enzymes actionand they can be detected in precooked, pasteurizedand manufactured foods. The direct detection ofenterotoxins in foods requires development ofpractical, rapid and sensitive assays. Currently, thebest methods for enterotoxin identification andquantification depend on the availability of specificantibodies for each enterotoxin type.Immunological methods with monoclonal andpolyclonal antibodies for the enterotoxin detectionat the level of 1 - 2 ng g