Characterization of a Broadly Specific LPS-Neutralizing Single Domain Antibody

Authors

  • V. Yadav
  • A. Singh Professor (Immunology), Department of Veterinary Microbiology, Lala Lajpat Rai University of Veterinary & Animal Sciences, Hisar, Haryana India
  • J. Rawat

DOI:

https://doi.org/10.37591/rrjoi.v7i1.1183

Abstract

Gram negative endotoxaemia or sepsis remains a lethal challenge to humans and animals, globally. Single domain antibodies (dAbs) have emerged as therapeutic biologics for challenging antigens such as lipopolysachharide (LPS). The present study characterized a broadly specific LPS-neutralising dAb produced from phage display library from LPS-immunized Indian desert camel, for its sequence validation with mass spectrometry. Also, the dAb clone 26 was subjected to co-crystallisation with LPS, extracted from E. coli O6 (ATCC 25922). The mass spectrometry of dAb clone 26 was performed upon in-gel trypsin digestion, MALDI-TOF analysis of the derived peptides and sequence identification on Mascot protein identification. The co-crystallization grid screening was performed with affinity purified dAb clone 26 in complex with LPS in a hanging drop vapor diffusion set up with pH and various precipitants i.e., ammonium sulphate, PEG3350, 6000 and 8000 as variables. The sequence of dAb clone 26 was validated with the sequence coverage of 35% through mass spectrometry. The study led to further validation of dAb clone 26 for developing it as a therapeutic candidate for endotoxemia.

 

Keywords: Endotoxin, single domain antibody, MALDI-TOF, dAb-LPS co-crystallization

 

Cite this Article

Yadav V, Singh A, Rawat J. Characterization of a Broadly Specific LPS-Neutralizing Single Domain Antibody. Research & Reviews: Journal of Immunology. 2017; 7(1): 1–8p.

Downloads

Published

2019-04-02

Issue

Vol. 7 No. 1 (2017)

Section

Research Article

License

Declaration and Copyright Transfer Form

(to be completed by authors)

I/ We, the undersigned author(s) of the submitted manuscript, hereby declare, that the above manuscript which is submitted for publication in the STM Journals(s), is not published already in part or whole (except in the form of abstract) in any journal or magazine for private or public circulation, and, is not under consideration of publication elsewhere.

  • I/We will not withdraw the manuscript after 1 week of submission as I have read the Author Guidelines and will adhere to the guidelines.
  • I/We Author(s ) have niether given nor will give this manuscript elsewhere for publishing after submitting in STM Journal(s).
  • I/ We have read the original version of the manuscript and am/ are responsible for the thought contents embodied in it. The work dealt in the manuscript is my/ our own, and my/ our individual contribution to this work is significant enough to qualify for authorship.
  •  I/We also agree to the authorship of the article in the following order:

Author’s name                                    

 

1. ________________

2. ________________

3. ________________

4. ________________

 

We  Author(s) tick this box and would request you to consider it as our signature as we agree to the terms of this Copyright Notice, which will apply to this submission if and when it is published by this journal.