Development and validation of an immunoassay for quantification of NCAM-1 in human plasma

Arcan Guven; Kayleigh Gray; Kuan-Wei Peng; Allison Klotz; Mark D Kellogg; Niven R Narain; Michael A Kiebish

doi:10.1016/j.jpba.2021.113981

Development and validation of an immunoassay for quantification of NCAM-1 in human plasma

J Pharm Biomed Anal. 2021 Apr 15:197:113981. doi: 10.1016/j.jpba.2021.113981. Epub 2021 Feb 16.

Authors

Arcan Guven¹, Kayleigh Gray², Kuan-Wei Peng², Allison Klotz², Mark D Kellogg³, Niven R Narain², Michael A Kiebish²

Affiliations

¹ BERG, 500 Old Connecticut Path, Framingham, MA, 01701, USA. Electronic address: arcan.guven@berghealth.com.
² BERG, 500 Old Connecticut Path, Framingham, MA, 01701, USA.
³ BERG, 500 Old Connecticut Path, Framingham, MA, 01701, USA; Department of Pathology, Harvard Medical School, Boston MA and Department of Laboratory Medicine, Boston Children's Hospital, 300 Longwood Avenue, Boston, MA, 02115, USA.

PMID: 33657526
DOI: 10.1016/j.jpba.2021.113981

Abstract

Background: Neural Cell Adhesion Molecule 1 (NCAM-1), a multifunctional member of the immunoglobulin superfamily, is expressed on the surface of neurons, glia, skeletal muscle, and natural killer cells. NCAM-1 has been implicated as having a role in cell-cell adhesion, involved in development of the nervous system, and for cells involved in the expansion of T cells and dendritic cells which play an important role in immune surveillance. Sensitive and specific methods to quantify non-surface bound NCAM-1 are not available.

Method: A sandwich ligand binding assay was developed for quantification of NCAM-1 in plasma and validated using an electro-chemiluminescent (ECL) technology.

Results: The data presented here demonstrated that the validated method met all prespecified criteria for precision, linearity, and accuracy in the range of 62.5 ng/mL to 4000.0 ng/mL, the range believed to be most relevant for plasma. The bioanalytical validation of the assay established the inter-assay coefficient of variation <8 % for calibration points, <2 % for high quality control (HQC), <8 % for medium quality control (MQC) and <19 % for low quality control (LQC) samples. Purified NCAM-1 spike-recovery experiment in plasma was used to determine assay accuracy; nominal concentrations (%) of NCAM-1 ranged from 91 % to 112 % for high and low spike level, respectively. Assay performance was subsequently evaluated for parallelism, selectivity, interference, and stability.

Conclusion: NCAM-1 assay has been developed and validated in human plasma and met all assay validation parameters pre-determined during development. Clinical testing of human plasma samples indicated that NCAM-1 does not seem to be influenced by age and was slightly influenced by gender. NCAM-1 assay has potential to be used as a biomarker assay once the assay is subjected to appropriate clinical assessment and diagnostic thresholds are established.

Keywords: Assay development; Assay validation; Biomarker; Immunoassay; Ligand binding assay; NCAM-1.

MeSH terms

Biomarkers
CD56 Antigen
Calibration
Humans
Immunoassay
Neural Cell Adhesion Molecules*
Quality Control

Substances

Biomarkers
CD56 Antigen
NCAM1 protein, human
Neural Cell Adhesion Molecules