Box contains

1. 96-well Protein Binding Plate (Part No. 231001): One strip well 96-well plate.

2. Anti-DNP Antibody (1000X) (Part No. 231002): One 20 µL vial of anti-DNP Rabbit IgG.

3. Secondary Antibody, HRP Conjugate (1000X) (Part No. 231009): One 20 µL vial.

4. 25X DNPH Solution (Part No. 231010): One 500 µL amber vial.

5. 2X DNPH Diluent (Part No. 231005): One 15 mL bottle.

6. Blocking Reagent (Part No. 231006): One 20 g bottle.

7. 10X Wash Buffer (Part No. 310806): One 100 mL bottle.

8. Substrate Solution (Part No. 310807): One 12 mL amber bottle.

9. Stop Solution (Part. No. 310808): One 12 mL bottle.

10. Reduced BSA Standard (Part No. 231007): One 200 µL vial of 1 mg/mL fully reduced BSA in

PBS.

11. Oxidized BSA Standard (Part No. 231008): One 200 µL vial of 1 mg/mL oxidized BSA in PBS at

7.5 nmol protein carbonyl/mg proteins. The protein carbonyl is predetermined by a

spectrophotometric method as described by Reznick and Parker (See Ref. 5).

Materials Not Supplied

1. Protein samples such as purified protein, plasma, serum, cell lysate, or tissue homogenate

2. 1X PBS

3. Ethanol

Storage

Upon receipt, aliquot and store both the Reduced and Oxidized BSA Standards at -20ºC to avoid

multiple freeze/thaw cycles. Store all other components at 4ºC.

Preparation of Reagents

• 1X Wash Buffer: Dilute the 10X Wash Buffer Concentrate to 1X with deionized water. Stir to

homogeneity.

• 1X DNPH Diluent: Dilute the 2X DNPH Diluent to 1X with deionized water. Mix well.

• Blocking Solution: Weigh out 5 g of Blocking Reagent, dissolve in 100 mL of 1X PBS, and store

at 4ºC for up to one week.

• DNPH Working Solution: Based on the number of tests, FRESHLY prepare appropriate amount of

DNPH Working Solution by diluting the 25X DNPH Solution to 1X in 1X DNPH Diluent. For

example: for 20 assays, transfer 80 μL of 25X DNPH Solution to a tube containing 1.92 mL of 1X

DNPH Diluent, mix well and use it IMMEDIATELY.

• Anti-DNP Antibody and Secondary Antibody: Immediately before use dilute the Anti-DNPH

antibody 1:1000 and Secondary Antibody 1:1000 with 1X Blocking Solution. Do not store diluted

solutions.