DESCRIPTION

USP36 (Ubiquitin Specific Peptidase 36) is a deubiquitinating enzyme, belonging to the USP family, that regulates protein stability and function mainly by removing ubiquitin chains from target proteins. USP36 plays a significant role in the nucleolus, where it stabilizes nucleolar proteins such as B23 and fibrillarin through deubiquitination, thereby regulating ribosome biogenesis. Additionally, USP36 is involved in the regulation of oxidative stress responses by stabilizing the mitochondrial superoxide dismutase SOD2. In terms of disease, USP36 is associated with the development and progression of various cancers. For instance, in breast cancer, USP36 stabilizes the estrogen receptor α (ERα) through deubiquitination, promoting tumor progression and leading to resistance to endocrine therapy.

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	2,350.6	111.7	4.8
2	8	626.8	16.9	2.7
3	8	310.8	13.4	4.3

Inter-assay Precision

Sample	n	Mean (pg/mL)	SD	CV%
1	16	2,397.8	113.7	4.7
2	16	627.5	24.8	4.0
3	16	305.1	13.3	4.4

RECOVERY

The recovery of human USP36 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	82	75-88
	1:32	89	81-93

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	87
	Range (%)	81-93
1:8	Average% of Expected	87
	Range (%)	86-89
1:16	Average% of Expected	109
	Range (%)	94-124

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Human USP36 Sandwich ELISA Kit produced at Sunlong Medical™.