CEL ELISA 试剂盒

1. 96-well Protein Binding Plate : One strip well 96-well plate.
2. Anti-CEL Antibody (1000X) : One 10 μL vial of anti-CEL antibody.
3. Secondary Antibody, HRP Conjugate (1000X) : One 20 μL vial.
4. Assay Diluent : One 50 mL bottle.
5. 10X Wash Buffer : One 100 mL bottle.
6. Substrate Solution : One 12 mL amber bottle.
7. Stop Solution (Part. No. 310808): One 12 mL bottle.

Box 2 (shipped on blue ice packs)

1. Protein samples such as purified protein, plasma, serum, cell lysate
2. 1X PBS
3. 10 μL to 1000 μL adjustable single channel micropipettes with disposable tips
4. 50 μL to 300 μL adjustable multichannel micropipette with disposable tips
5. Multichannel micropipette reservoir
6. Microplate reader capable of reading at 450 nm (620 nm as optional reference wave length) 3

• Provides rapid detection and quantitation of CEL protein adducts
• Perform up to 96 assays, including standard curve and unknown protein samples
• Will not cross react with CML protein adducts

CEL Conjugate Coated Plate: Note: The CEL Conjugate coated wells are not stable and should be used within 24 hrs after coating. Only coat the number of wells to be used immediately. 1. Immediately before use, prepare 1X Conjugate Diluent by diluting the 100X Conjugate Diluent in 1X PBS. Example: Add 50 μL to 4.95 mL of 1X PBS. 2. Immediately before use, prepare 1.0 μg/mL of CEL Conjugate by diluting the 1.0 mg/mL CEL Conjugate in 1X Conjugate Diluent. Example: Add 5 μL of 1.0 mg/mL CEL Conjugate to 4.995 mL of 1X Conjugate Diluent and mix well. 3. Add 100 μL of the 1 μg/mL CEL Conjugate to each well to be tested and incubate overnight at 4 °C. Remove the CEL Conjugate coating solution and wash twice with 1X PBS. Blot plate on paper towels to remove excess fluid. Add 200 μL of Assay Diluent to each well and block for 1 hr at room temperature on an orbital shaker. Transfer the plate to 4 °C and remove the Assay Diluent immediately before use. 1X Wash Buffer: Dilute the 10X Wash Buffer to 1X with deionized water. Stir to homogeneity. Anti-CEL Antibody and Secondary Antibody: Immediately before use, dilute the Anti-CEL antibody 1:1000 and Secondary Antibody 1:1000 with Assay Diluent. Do not store diluted solutions.

1. Prepare and mix all reagents thoroughly before use. Each CEL sample including unknown and standard should be assayed in duplicate.
2. Add 50 μL of the unknown sample or CEL-BSA standard to the wells of the CEL Conjugate coated plate. If needed, unknown samples may be diluted in 1X PBS containing 0.1 % BSA before adding. Incubate at room temperature for 10 minutes on an orbital shaker.
3. Add 50 μL of the diluted anti-CEL antibody to each well, incubate at room temperature for 1 hour on an orbital shaker.
4. Wash 3 times with 250 μL of 1X Wash Buffer with thorough aspiration between each wash. After the last wash, empty wells and tap microwell strips on absorbent pad or paper towel to remove excess 1X Wash Buffer.
5. Add 100 μL of the diluted Secondary Antibody-HRP Conjugate to all wells and incubate for 1 hour at room temperature on an orbital shaker. Wash the strip wells 3 times according to step 4 above.
6. Warm Substrate Solution to room temperature. Add 100 L of Substrate Solution to each well. Incubate at room temperature for 5-20 minutes on an orbital shaker. Note: Watch plate carefully, if color changes rapidly, the reaction may need to be stopped sooner to prevent saturation.
7. Stop the enzyme reaction by adding 100 μL of Stop Solution to each well. Results should be read immediately (color will fade over time).
8. Read absorbance of each well on a microplate reader using 450 nm as the primary wave length. 5