en Diagnostic/screening methods and protocols Diagnostic/screening methods and protocols Original article Original article <strong>Introduction:</strong> <em>Staphylococcus aureus</em> enterotoxin A (SEA) is one of the most common causes of staphylococcal food poisoning. Due to the simplicity and no requirement for laboratory apparatuses, dot-ELISA is a choice method for detecting Staphylococcal enterotoxins. The present study aimed to develop a dot-ELISA for the detection of SEA. <strong>Methods:</strong> Nitrocellulose membranes were coated with the SEA antibody and blocked by the addition of 3% bovine serum albumin (BSA) blocking buffer. After 1 h incubation and washing the membranes, milk samples and the positive control (SEA, 50 ng/ml) were added to the membranes and incubated for 1 h. The membranes were then washed and incubated for 45 min with HRP-conjugated SEA, followed by the addition of TMB. <strong>Results: </strong>Our dot-ELISA could detect amounts of &ge; 50 ng/ml of SEA in the milk samples. Of the 30 raw milk samples randomly purchased from dairy product stores in District 3, Tehran, 5 (16%) contained SEA &ge; 50 ng/ml by the dot-ELISA. <strong>Conclusion</strong>: The dot-ELISA showed to be a reliable method for the preliminary screening of milk samples for SEA contamination. This method is cost-effective, fast, and does not require an ELISA-reader device. dot-ELISA, Staphylococcus aureus Enterotoxin A (SEA), Raw milk 132 136 http://jommid.pasteur.ac.ir/browse.php?a_code=A-10-220-1&slc_lang=en&sid=1 Haniyeh Golafrouz haniyeh.golafrouz@yahoo.com 0000-0003-2648-5109 No Department of Agricultural Sciences and Food Technology, Science and Research Branch, Islamic Azad University, Tehran, Iran Hamed Ahari dr.h.ahari@gmail.com 0000-0002-4450-2454 Yes Department of Agricultural Sciences and Food Technology, Science and Research Branch, Islamic Azad University, Tehran, Iran Seyed Amirali Anvar saaa4824@gmail.com 0000-0002-6204-5621 No Faculty of Veterinary Medicine, Islamic Azad University, Food Hygiene Sciences, Tehran, Iran Delavar Shahbazzadeh shahbazzadeh@yahoo.com 0000-0002-9066-8666 No Pasteur Institute of Iran, Biotechnology Research Center, Venom and Bio therapeutic Molecules Lab., Tehran-Iran.