Cat. No.	K0331221
Product	Human RANTES ELISA Kit
Type	ELISA Kit
Quantity	96-well assay
Specificity	Human RANTES ELISA kit contains all components required for the quantitative measurement of natural and/or recombinant Human RANTES in serum or other biological samples in a sandwich ELISA format within the range of 32-3000 pg/ml.
Description	Using the ELISA protocol described below, the components supplied in this kit are sufficient to assay Human RANTES in approximately 96 ELISA plate wells.
Kit Components	Antigen-affinity purified rabbit anti-Human RANTES Pre-Coated 96 well ELISA microplate Detection Antibody : Biotinylated antigen-affinity purified rabbit anti-Human RANTES Standard Protein (Lyophilized) : Recombinant Human RANTES Color development Enzyme (Lyophilized) : Avidin-HRP conjugate Assay Diluting Solution : 0.1% BSA in PBST (50 ml) Color development Reagent A : TMB solution (10 ml) Color development Reagent B : H2O2 solution (20 ml) Stop Solution : 2M H2SO4 (10 ml) PBS powder Tween-20 (50%)
Dinamic Range	32pg/ml to 3000 pg/ml
Cross Reactivity	Human
Storage & Shelf life	1 year at 2-8°C
Reconstitution	1. Human RANTES Standard: 100ng (1 vial) of recombinant Human RANTES should be reconstituted in 100 ul sterile water for a concentration of 1ug/ml. 2. Detection Antibody: 2.5 ug (1 vial) of biotinylated antigen- affinity purified anti-Human RANTES should be reconstituted in 0.25 ml sterile water for a concentration of 10 ug/ml.
Buffer Preparation	1. Washing solution (PBST): Resolve the PBS powder (1 pouch) to sterile water and make 1Liter, then add 50% 1ml Tween-20 to this solution and mix well. 2. Pre-coated ELISA well plate: Select the number of coated wells required for the assay. The remaining wells should be placed in the resealable pouch with a desiccant. The pouch must be resealed to protect from moisture. 3. Sample and Standard dilution: Dilute the reconstituted standard from 4 ng/ml to zero in assay diluting solution. Dilute the samples to a proper concentration in assay diluting solution. Each well needs 100 ul. 4. Detection Antibody: Dilute the reconstituted detection antibody in assay diluting solution to a concentration of 0.25ug/ml (1/40 dilution). Each well needs 100 ul. 5. Color development enzyme: Dilute the avidin-HRP conjugate 1:2000 in assay diluting solution. Each well needs 100 ul. 6. Color development solution: Mix 1 volume of color development reagent A and 2 volume of reagent B (1:2) prior to use. Each well needs 100 ul.
Sandwich ELISA Protocol	1. Add 200 ul of PBS (or DW) to each well. Aspirate the wells to remove liquid and wash the plate 3 times using 300 ul of washing solution per well. After the last wash invert plate to remove residual solution and blot on paper towel. 2. Add 100 ul of standard or sample to each well in duplicate. Incubate at room temperature for at least 2 hours. (or 37°C for 1hour) 3. Aspirate the wells to remove liquid and wash the plate 4 times like as step 1. 4. Add 100 ul of the diluted detection antibody (0.25 ug/ml) per well. Incubate at room temperature for 2 hours. (or 37°C for 1 hour) 5. Aspirate and wash plate 4 times like as step 1. 6. Add 100 ul of the diluted color development enzyme (1/200 dilute) per well. Incubate 30 minutes at room temperature. (or 37°C for 30 minutes) 7. Aspirate and wash plate 4 times like as step 1. 8. Add 100 ul of color development solution to each well. Incubate at room temperature for a proper color development. (5-30 minutes) To stop the color reaction, add 50 ul of the stop solution to each well. 9. Using a microtiter plate reader, read the plate at 450 nm wavelength.

RANTES ELISA Kit, human