DESCRIPTION

CD6 is a lymphocyte surface accessory molecule that is associated with the T-cell receptor (TCR) complex in the immune synapse. It plays a key role in T-cell activation and proliferation by mediating cell-cell contacts and regulating T-cell responses. CD6 is involved in the stabilization and maturation of the immunological synapse, facilitating T-cell interactions with antigenpresenting cells (APCs) and subsequent T-cell proliferative responses. Additionally, CD6 has been shown to modulate T-cell activation by attenuating signaling cascades, acting as a rheostat-type signalosome that tunes T-cell activation.

PRINCIPLE OF THE ASSAY

An anti-tag antibody is pre-coated onto the bottom of wells. After adding antigen or samples, Tagged-Capture antibody and HRP-conjugated detection antibody, a sandwich complex is formed in the solution. TMB acts as a HRP substrate, and the solution color will change from colorless to blue. A stop solution containing sulfuric acid turns the 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 (pg/mL)	SD	CV%
1	8	2,693.5	41.7	1.5
2	8	643.1	7.3	1.1
3	8	324.4	5.5	1.7

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	16	2,688.3	47.4	1.8
2	16	649.7	12.7	2.0
3	16	325.0	5.4	1.7

RECOVERY

The recovery of human CD6 spiked to three different levels throughout the range of the assay in various matrices was evaluated.

Sample Type		Average% of Expected	Range (%)
Human serum	1:16	119	111-126
	1:32	120	116-122
Cell lysate	1:64	79	76-83
	1:128	81	77-83

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. (The human serum was initially diluted 1:2. The cell lysate was initially diluted 1:8.)

		Human serum	Cell lysate
1:2	Average% of Expected	100	100
	Range (%)	-	-
1:4	Average% of Expected	110	96
	Range (%)	107-114	93-99
1:8	Average% of Expected	115	108
	Range (%)	108-122	108-109
1:16	Average% of Expected	110	124
	Range (%)	96-124	119-130

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human CD6 produced at Sunlong Medical™.