DESCRIPTION

STK39 (Serine/Threonine Kinase 39) is a prolineand alanine-rich Ste20-related kinase (also named as SPAK), which is composed of a short N-terminal proline and alanine repeats (PAPA box), a kinase catalytic domain and a C-terminal regulatory domain. In mammalians, STK39 plays an important role in ion homeostasis by regulating the cation chloride cotransporters' activities, which is crucial for the modulation of renal salt transport and blood pressure. Notably, STK39 is also implicated in the regulation of tumor cell proliferation, migration, and invasion in multiple cancers, including osteosarcoma and cervical cancer.

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	2,509.6	118.6	4.7
2	8	507.3	16.7	3.3
3	8	254.0	9.6	3.8

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	16	2,443.7	138.3	5.7
2	16	510.3	15.0	2.9
3	16	260.9	12.2	4.7

RECOVERY

The recovery of human STK39 spiked to three different levels throughout the range of the assay in cell lysate was evaluated.

Sample Type		Average% of Expected	Range (%)
Cell lysate	1:16	96	78-104

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 1:2.)

		Cell lysate
1:2	Average% of Expected	100
	Range (%)	-
1:4	Average% of Expected	104
	Range (%)	103-104
1:8	Average% of Expected	106
	Range (%)	98-113
1:16	Average% of Expected	100
	Range (%)	82-117

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Human STK39 Sandwich ELISA Kit produced at Sunlong Medical™.