symmacros
第6楼2009/03/29
供参考
ELISA method for Fumonsins
Procedure (ELISA)
1. extract with methanol:water (70:30)
2a. dilute 10X with binding buffer no conjugate (confirmation)
2b. dilute 10X with binding buffer/conjugate (quantitation)
3. add to well, mix for 20 minutes
3. wash five times with distilled water
5a. add 100% methanol and remove (MS confirmation)
5b. add substrate solution (TMB-peroxide)
6. after 10 minutes add dilute sulfuric acid
LC/Fluorescence (OPA)
sample preparation
1.extract sample (25g) with methanol:water (70:30)
2.dilute 1:4 with water
3.apply extract (60%) to C18 or SAX column
4.wash and elute
5.evaporate elute to dryness
6.dissaolve residue in 50:50 water:acetonitrile
7.add OPA reagent, mix and analyze
Experimental
Fluorescence detector (ex.335, em.440)
C18 column (4.6 X 150mm, 3um particles)
Flow rate: 1ml/min
Mobile phase: 48:52 water:acetonitrile
1% acetic acid in mobile phase