DESCRIPTION

Used for the quantitative determination of Anti-Atezolizumab concentration in serum and plasma.

PRINCIPLE OF THE ASSAY

This assay employs the quantitative sandwich enzyme immunoassay technique. Atezolizumab has been pre-coated onto a microplate. Samples or standards are pipetted into microwells and Anti-Atezolizumab will be captured by immobilized Atezolizumab. After washing away any unbound substances, a biotin-labeled Atezolizumab is added to the wells. After washing away any unbound substances, Streptavidin-HRP is added to the wells. Following a wash to remove any unbound enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of Anti-Atezolizumab bound in the initial step. The color development is stopped and the intensity of the color is measured.

PRECISION

Intra-Assay Precision (Precision within an assay): <10% Three samples of known concentration were tested sixteen times on one plate to assess intra-assay precision. Inter-Assay Precision (Precision between assays): <15% Three samples of known concentration were tested in twenty four separate assays to assess inter-assay precision.

Sample		1	2	3	1	2	3
n		16	16	16	24	24	24
Mean (ng/mL)		47.6	11.8	2.5	49.0	11.7	3.1
Standard deviation		1.3	0.5	0.1	1.8	0.4	0.2
CV \(%\)		2.8	4.4	5.6	3.7	3.6	7.0

RECOVERY

The recovery of Anti-Atezolizumab spiked to three different levels in human serum samples throughout the range of the assay was evaluated.

Sample	Dilution factor	Average (%)	Range (%)
Human serum (n=4)	1:10	93	82-108
	1:20	93	83-103

LINEARITY

To assess the linearity of the assay, human serum samples were spiked with high concentrations of Anti-Atezolizumab and diluted with the Standard Diluent to produce samples with values within the dynamic range of the assay.

Sample	Dilution factor	Average (%)	Range (%)
Human serum (n=4)	1:2	96	93-98
	1:4	96	93-100
	1:8	99	83-115
	1:16	99	87-110