检测大肠杆菌试纸条的制作资料.ppt

1、The cultivation of EHEC 0157: H7 2、McAb epitopes identification 2.1 Determination of the optimal antigen coating concentration The antigen ：1x108、5x107、2.5x107、1.25x107、5x106 2.5x106、1.25x106、5x105 Coated on ELISA plates 4°C 12h Monoclonal antibodies ：1H1、1H3、2A2、3C5、4B10、 4E4、4H10、6F4 2、McAb epitopes identification 2.2 Identification of McAb epitopes： coat seal cell supernatant secondary antibody developing termination count 3.Hybridoma recovery and McAb preparation 3.1 Hybridoma recovery 3.2 McAb ascites production and purification 4、Preparation of colloidal gold test strip for detecting EHEC 0157:H7 4.1preparation of the colloidal gold 4.2 optimal amount of McAb for the conjugation with colloidal gold 4.3 optimal pH for the conjugation between colloidal gold and McAb 4.4 Colloidal gold mark McAb preparation and purification  4.5.1 Coating quantity l0μL/cm、15μL/cm、20μL/cm vacuum drying 4h 4.5.2 The choice of block solution 4.5.3 The choice of the sample diluent 0.0lmol/L PBS、0.0lmol/L PBS +0.05%Triton. 0.0lmol/L PBS + 0.05%Tween、0.0lmol/L PBS +20% 0.5mol/L Tris-HCl+0.05%Triton pH 6.0、6.5、7.0、7.5、8.0、8.5、9.0 5 The colloidal gold test strip performance evaluation 5.1 Specificity TSB 37℃ 12h 100℃ 30min 107CFU/mL-108CFU/mL 5.2 sensitivity Ixl08CFU/mL、Ixl07CFU/mLlxl06CFU/mL、lxl05CFU/mL lxl04CFU/mL Ixl03CFU/mL 5.3 repeatability 4℃ 0d、3d、6d、9d 6 Market pork sample detection conventional method * Development of Colloidal Gold Test Strip for Detection of EHEC O157:H7 1、The cultivation of EHEC 0157: H7 2、McAb epitopes identification 3、Hybridoma recovery and McAb preparation 4、Preparation of Colloidal Gold Test Strip for Detection of EHEC O157:H7 5、Colloidal gold test strip performance evaluation 6、Market pork sample detection EHEC O157:H7 CT-SMAC 37℃ 18h mEC+n 37℃ 12h colony co