ELISA Development Guide - OriGene

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ev022912 OriGene Technologies, Inc. 9620 Medical Center Drive, Suite 200, Rockville, MD 20850 www.origene.com techsupport@origene.com 1-301-340-3188 (P) 1-301-340-9254 (F) Page 2
ev022912 Table of Contents ELISA Plate Coating .....................................................................................................................4 Materials .................................................................................................................................4 Antibody Plate Coating Procedure ...........................................................................................5 Basic ELISA Protocol ...................................................................................................................6 Materials .................................................................................................................................6 Recommended Plate Layout ....................................................................................................7 Preparation of Materials and Buffers ......................................................................................7 Detailed ELISA Assay Protocol .................................................................................................8 Data Analysis .........................................................................................................................10 ELISA Protocol Summary .......................................................................................................11 ELISA Assay Optimization .........................................................................................................12 ELISA Assay Troubleshooting ....................................................................................................15 Preparing Antibody-Coupled Luminex Beads ...........................................................................19 Materials and Equipment Required .......................................................................................19 Set-Up ...................................................................................................................................20 Procedure Notes ...................................................................................................................20 Coupling Procedure ...............................................................................................................21 Luminex Assay Protocol............................................................................................................23 Required or Recommended Reagents and Supplies ...............................................................23 Equipment and software required: ........................................................................................24 Setting the Luminex 100 or 200 for “High Gain” or “High PMT” Reading ...............................25 Procedure Notes ...................................................................................................................25 Recommended Plate Layout ..................................................................................................26 Sample Collection and Preparation .......................................................................................26 Dilution of Samples ...............................................................................................................27 Reagent Preparation .............................................................................................................27 Assay Protocol .......................................................................................................................28 Data Analysis .........................................................................................................................31 Luminex ELISA Protocol Summary ............................................................................................32 Luminex Assay Optimization ....................................................................................................33 Luminex ELISA Troubleshooting ...............................................................................................36 Buffer and Reagent Preparation ...............................................................................................38 Plate Plan Template .................................................................................................................41 OriGene Technologies, Inc. 9620 Medical Center Drive, Suite 200, Rockville, MD 20850 www.origene.com techsupport@origene.com 1-301-340-3188 (P) 1-301-340-9254 (F) Page 3
Page 1: ev022912 ELISA Development Guide A
Page 5 and 6: ev022912 Storage of antibody-coated
Page 7 and 8: ev022912 Recommended Plate Layout B
Page 9 and 10: ev022912 d. Assay uses 48 wells. e.
Page 11 and 12: ev022912 ELISA Protocol Summary 1.
Page 13 and 14: ev022912 Parameter Modification Pos
Page 15 and 16: ev022912 ELISA Assay Troubleshootin
Page 17 and 18: ev022912 Problem Possible Causes So
Page 19 and 20: ev022912 Preparing Antibody-Coupled
Page 21 and 22: ev022912 Coupling Procedure Step 1
Page 23 and 24: ev022912 Luminex Assay Protocol Req
Page 25 and 26: ev022912 Note : To achieve optimal
Page 27 and 28: ev022912 7. For accurate quantitati
Page 29 and 30: ev022912 7. Ten to fifteen minutes
Page 31 and 32: ev022912 30. Optional: shake plate
Page 33 and 34: ev022912 Luminex Assay Optimization
Page 35 and 36: ev022912 Parameter Modification Pos
Page 37 and 38: ev022912 Problem Cause Solution Hig
Page 39 and 40: ev022912 Post-Coat/Blocking Buffer
Page 41: ev022912 Plate Plan Template 12 11

ELISA Development Guide - OriGene

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