A chemiluminescent-western blot assay for quantitative detection of Plasmodium falciparum circumsporozoite protein

Sanjai Kumar, Hong Zheng, Davison T. Sangweme, Babita Mahajan, Yukiko Kozakai, Phuong T. Pham, Merribeth J. Morin, Emily Locke, Nirbhay Kumar

Research output: Contribution to journalArticle

Abstract

Highly sensitive and reliable assays based on the quantitation of immunologically relevant component(s) in recombinant or whole parasite-based vaccines would facilitate pre-clinical and clinical phases and the monitoring of malaria vaccine deployment. Here we report a laboratory-grade Western Blot assay for quantitative detection of Plasmodium falciparum circumsporozoite protein (PfCSP) in P. falciparum sporozoite (PfSPZ) and in recombinant (rPfCSP) product. This assay is based on the immuno-reactivity of an anti-P. falciparum CSP monoclonal antibody (mAb 2A10) with the NANP-repeat units on PfCSP. The antigen-antibody complex is detected by reaction with a commercially obtained chemiluminescence-linked Immunodetection system. The linear range for detecting the recombinant P. falciparum CSP (rPfCSP) in this assay is 3-12pg (R2=0.9399). The range for detecting the day 15 salivary-gland PfSPZ is between 0.0625 and 1 parasite (R2=0.9448) and approximately 10.0pg of PfCSP was detected on each sporozoite. The assay was highly reproducible in measuring the PfCSP on PfSPZ. The inter-assay Coefficient of Variation (CV%) was 10.31% while the intra-assay CV% on three different days was 6.05%, 2.03% and 1.42% respectively. These results suggest that this ECL-WB assay is highly sensitive and robust with a low degree of inter-assay and intra-assay variations. To our knowledge, this is the most sensitive immunoassay for the detection of a recombinant or native malarial protein and may have a wider range of applications including the quantification of immunological component(s) in a vaccine formulation, determination of the antigenic integrity in adjuvanted-vaccine and in stability studies. In addition, this assay can be applied to measure the mosquito infectivity in malaria transmission areas and to determine the effects of intervention measures on malaria transmission.

Original languageEnglish (US)
Pages (from-to)99-105
Number of pages7
JournalJournal of Immunological Methods
Volume390
Issue number1-2
DOIs
StatePublished - Apr 30 2013

Fingerprint

Plasmodium falciparum
Western Blotting
Sporozoites
Malaria
Parasites
Vaccines
Vaccine Potency
Protozoan circumsporozoite protein
Malaria Vaccines
Luminescence
Antigen-Antibody Complex
Salivary Glands
Culicidae
Immunoassay
Monoclonal Antibodies

Keywords

  • Chemiluminescent-western blot
  • Circumsporozoite protein
  • IOD
  • Plasmodium falciparum

ASJC Scopus subject areas

  • Immunology
  • Immunology and Allergy

Cite this

A chemiluminescent-western blot assay for quantitative detection of Plasmodium falciparum circumsporozoite protein. / Kumar, Sanjai; Zheng, Hong; Sangweme, Davison T.; Mahajan, Babita; Kozakai, Yukiko; Pham, Phuong T.; Morin, Merribeth J.; Locke, Emily; Kumar, Nirbhay.

In: Journal of Immunological Methods, Vol. 390, No. 1-2, 30.04.2013, p. 99-105.

Research output: Contribution to journalArticle

Kumar, S, Zheng, H, Sangweme, DT, Mahajan, B, Kozakai, Y, Pham, PT, Morin, MJ, Locke, E & Kumar, N 2013, 'A chemiluminescent-western blot assay for quantitative detection of Plasmodium falciparum circumsporozoite protein', Journal of Immunological Methods, vol. 390, no. 1-2, pp. 99-105. https://doi.org/10.1016/j.jim.2013.02.001
Kumar, Sanjai ; Zheng, Hong ; Sangweme, Davison T. ; Mahajan, Babita ; Kozakai, Yukiko ; Pham, Phuong T. ; Morin, Merribeth J. ; Locke, Emily ; Kumar, Nirbhay. / A chemiluminescent-western blot assay for quantitative detection of Plasmodium falciparum circumsporozoite protein. In: Journal of Immunological Methods. 2013 ; Vol. 390, No. 1-2. pp. 99-105.
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