DESCRIPTION

TMEM16A/DOG1 gene encodes a member of Ca2+ activated Clchannels (CaCCs) that are critical for physiological functions, such as epithelial secretion, smooth muscle contraction and sensory signal transduction. TMEM16A/DOG1 has been reported to be a valuable target in noncancerous diseases. In cancer, TMEM16A/DOG1 was originally recognized to be positively expressed in gastrointestinal stromal tumor (GIST) and was more well‐recognized as DOG1 (Discovered On GISTs protein 1).

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 (pg/mL)	SD	CV%
1	8	113.8	4.4	3.9
2	8	31.0	2.2	7.1
3	8	17.2	0.8	4.4

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	16	117.5	7.0	5.9
2	16	30.6	2.0	6.6
3	16	16.5	1.0	6.0

RECOVERY

The recovery of human TMEM16A/DOG1 spiked to three different levels throughout the range of the assay in cell lysate was evaluated.

Sample Type		Average% of Expected	Range (%)
Cell lysate	1:80	107	82-122

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 cell lysate was initially diluted 1:10.)

		Cell lysate
1:2	Average% of Expected	100
	Range (%)	-
1:4	Average% of Expected	105
	Range (%)	93-116
1:8	Average% of Expected	93
	Range (%)	83-102
1:16	Average% of Expected	73
	Range (%)	70-77

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Human TMEM16A/DOG1 Sandwich ELISA Kit produced at Sunlong Medical™.