DESCRIPTION

Small GTPase Rab21 is a ubiquitously expressed and has recently been shown to associate with the α tails of several integrins via the shared conserved membrane proximal sequence, thus regulating cell adhesion and migration via controlling the endo/exocytic trafficking of most integrin heterodimers. Rab21 is required for the pruning of the immature neurites during the multipolar-to-bipolar transition. Knock down of Rab21 impairs integrin-mediated cell adhesion and motility, whereas its overexpression stimulates cell migration and cancer cell adhesion to collagen and human bone.

PRINCIPLE OF THE ASSAY

Sandwich ELISA structure (Detection antibody labeled with HRP) A capture antibody is pre-coated onto the bottom of wells which binds to analyte of interest. A detection antibody labeled with HRP also binds to the analyte. TMB acts as the HRP substrate and the solution color will change from colorless to blue. A stop solution containing sulfuric acid turns solution yellow. The color intensity is proportional to the quantity of bound protein which is measurable at 450 nm with the correction wavelength set at 630 nm.

PRECISION

Intra-assay Precision (Precision within an assay) Three samples of known concentration were tested 8 times on one plate to assess intra-assay precision. Inter-assay Precision (Precision between assays) Three samples of known concentration were tested in 16 separate assays to assess inter-assay precision.

Intra-assay Precision

Sample	n	Mean (ng/mL)	SD	CV%
1	8	5.64	0.53	9.37
2	8	1.35	0.04	2.93
3	8	0.68	0.03	4.01

Inter-assay Precision

Sample	n	Mean (ng/mL)	SD	CV%
1	16	5.19	0.29	5.50
2	16	1.20	0.16	13.23
3	16	0.60	0.08	13.34

RECOVERY

The recovery of human RAB21 spiked to three different levels throughout the range of the assay in cell lysate was evaluated.

Sample Type		Average% of Expected	Range (%)
Cell lysate	1:8	96	88-102
	1:16	102	99-107

LINEARITY

To assess the linearity of the assay, samples were diluted with the appropriate Sample Diluent to produce samples with values within the dynamic range of the assay.

		Cell lysate
1:2	Average% of Expected	100
	Range (%)	-
1:4	Average% of Expected	84
	Range (%)	80-89
1:8	Average% of Expected	92
	Range (%)	78-103
1:16	Average% of Expected	103
	Range (%)	92-121

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Human RAB21 Sandwich ELISA Kit produced at Sunlong Medical™.