This　work　reports　on　a　sensitive　colorimetric　immunoassay　for　aflatoxin　B1　(AFB1)　in　foodstuff.　The　reagent　L-ascorbic　acid　2-phosphate　(AAP)　is　added　to　the　system,　and　alkaline　phosphatase　(ALP)　hydrolyzes　AAP　under　formation　of　ascorbic　acid　and　phosphate.　The　ascorbic　acid　produced　reduces　chloroauric　acid　to　form　zero-valent　gold　(in　the　form　of　nanoparticles).　Hence,　Au(III)　is　no　longer　available　to　oxidize　2,2′-azinobis(3-ethylbenzthiazoline-6-sulfonate)　(ABTS)　to　form　a　green　product.　Rather,　the　solution　remains　colorless.　By　using　ALP-labelled　monoclonal　anti-AFB1　antibody,　a　competitive　enzyme-label　immunoassay　was　developed　for　AFB1　in　a　microplate　coated　with　the　AFB1-BSA　conjugate.　Under　optimal　conditions,　the　absorbance　of　the　solution　at　415 nm　increases　linearly　with　increasing　AFB1　concentration　in　range　from　10 pg·mL−1　to　100 ng·mL−1　(while　the　color　gradually　turns　to　green),　and　the　detection　limit　is　7.8 pg·mL−1.　The　precision　of　the　method　(expressed　as　RSD)　is　±9.7%.　The　accuracy　was　validated　by　analyzing　both　naturally　contaminated　and　spiked　food　samples,　and　the　results　matched　the　results　obtained　by　ELISA　very　well.　©　2017,　Springer-Verlag　Wien.