DESCRIPTION

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

PRINCIPLE OF THE ASSAY

This assay employs the quantitative sandwich enzyme immunoassay technique. Daratumumab has been pre-coated onto a microplate. Samples or standards are pipetted into microwells and Anti-Daratumumab will be captured by immobilized Daratumumab. After washing away any unbound substances, a biotin-labeled Daratumumab 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-Daratumumab 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)		833.0	201.1	85.2	835.9	210.9	89.5
Standard deviation		33.1	11.1	8.8	37.7	16.9	10.3
CV \(%\)		4.0	5.5	10.4	4.5	8.0	11.5

RECOVERY

The recovery of Anti-Daratumumab 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=3)	1:10	91	82-106
	1:20	88	81-97

LINEARITY

To assess the linearity of the assay, human serum samples were spiked with high concentrations of Anti-Daratumumab 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=5)	1:2	98	92-105
	1:4	102	93-112
	1:8	109	100-116
	1:16	104	95-113