DESCRIPTION

Used for the quantitative determination of Dupilumab concentration in serum and plasma.

PRINCIPLE OF THE ASSAY

This assay employs the quantitative sandwich enzyme immunoassay technique. An antibody specific for Dupilumab has been pre-coated onto a microplate. Standards or samples are pipetted into the wells and any Dupilumab present is bound by the immobilized antibody. After washing away any unbound substances, a biotin-labeled antibody specific for Dupilumab 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 Dupilumab 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
Standard deviation		99.8	47.4	13.1	130.0	85.2	14.7
CV \(%\)		4.4	9.4	9.3	6.0	14.5	11.2

RECOVERY

The recovery of Dupilumab 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	90	81-107
	1:20	87	80-91

LINEARITY

To assess the linearity of the assay, human serum samples were spiked with high concentrations of Dupilumab 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=3)	1:2	103	98-109
	1:4	101	90-107
	1:8	92	80-102
	1:16	94	82-110