The Human IP-10 Standard has been tested to assure to function as a standard when used with the BD™ Cytometric Bead Array (CBA), such as the BD™ CBA Human IP-10 Flex Set (Cat. No. 558280). Investigators are encouraged to refer to the technical data sheet for the BD™ CBA Human IP-10 Flex Set (Cat. No. 558280). An abbreviated instruction has been provided below (Preparation of Human IP-10 Standard Protocol). The Human IP-10 Standard should be reconstituted using Assay Diluent, which is provided with the BD™ CBA Human Soluble Protein Master Buffer Kit (Cat. No. 558264 or 558265) or it may also be purchased separately (Cat. No. 560104).
Preparation of Human IP-10 Standard: The Human IP-10 Standard is lyophilized and should be reconstituted and serially diluted before mixing with CBA capture beads and PE detection reagents.
1. Open one vial of the lyophilized Human IP-10 Standard. Transfer the lyosphere(s) to a polypropylene tube (BD Falcon™, Cat. No. 352097). Label the tube "Top Standard".
2. Reconstitute the standard with 4.0 mL of Assay Diluent. Allow the reconstituted standard to equilibrate for at least 15 minutes before making dilutions. Mix the reconstituted protein by pipette only. Do not vortex or mix vigorously.
3. Label additional 12 x 75 mm tubes (BD Falcon™, Cat. No. 352008) and arrange them in the following order: Top Standard, 1:2, 1:4, 1:8, 1:16, 1:32, 1:64, 1:128, and 1:256.
4. Add 500 µl of Assay Diluent to each of the dilution tubes.
5. Perform a serial dilution by transferring 500 µl from the Top Standard to the 1:2 dilution tube and mix thoroughly. Do not vortex, mix by pipet only. Continue making serial dilutions by transferring 500 µl from the 1:2 tube to the 1:4 tube and so on to the 1:256 tube and mix thoroughly. Prepare one tube containing only Assay Diluent to serve as the 0 pg/mL negative control.