A novel approach for the chromatographic purification and peptide mass fingerprinting of urinary free light chains

[Display omitted] •Urinary free light chains (λ-FLCs and κ-FLCs) have been purified.•Detection of urinary FLCs was achieved by immunoblotting using FLC specific monoclonal antibody.•Peptide mass fingerprint analysis of λ-FLCs and κ-FLCs has yielded peptides that partially match with gi|218783338 and...

Full description

Saved in:
Bibliographic Details
Published in:International journal of biological macromolecules 2017-02, Vol.95, p.331-339
Main Authors: Mali, Bhupesh C., Badgujar, Shamkant B., Shukla, Kunal K., Bhanushali, Paresh B.
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Bence Jones proteins
Chromatography - methods
Free light chains
Humans
Immunoblotting
Immunoglobulin kappa-Chains - chemistry
Immunoglobulin kappa-Chains - isolation & purification
Immunoglobulin kappa-Chains - urine
Immunoglobulin lambda-Chains - chemistry
Immunoglobulin lambda-Chains - isolation & purification
Immunoglobulin lambda-Chains - urine
Medical Waste
Peptide Mapping - methods
Proteomics
Urine
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:[Display omitted] •Urinary free light chains (λ-FLCs and κ-FLCs) have been purified.•Detection of urinary FLCs was achieved by immunoblotting using FLC specific monoclonal antibody.•Peptide mass fingerprint analysis of λ-FLCs and κ-FLCs has yielded peptides that partially match with gi|218783338 and gi|48475432 respectively.•This is the first record on characterization and validation of Urinary FLCs. We describe a chromatographic approach for the purification of urinary free light chains (FLCs) viz., lambda free light chains (λ-FLCs) and kappa free light chains (κ-FLCs). Isolated urinary FLCs were analyzed by SDS-PAGE, immunoblotting and mass spectrometry (MS). The relative molecular masses of λ-FLC and κ-FLC are 22,933.397 and 23,544.336Da respectively. Moreover, dimer forms of each FLC were also detected in mass spectrum which corresponds to 45,737.747 and 47,348.028Da respectively for λ-FLCs and κ-FLCs. Peptide mass fingerprint analysis of the purified λ-FLCs and κ-FLCs has yielded peptides that partially match with known light chain sequences viz., gi|218783338 and gi|48475432 respectively. The tryptic digestion profile of isolated FLCs infers the exclusive nature of them and they may be additive molecules in the dictionary of urinary proteins. This is the first report of characterization and validation of FLCs from large volume samples by peptide sequencing. This simple and cost-effective approach to purification of FLCs, together with the easy availability of urine samples make the large-scale production of FLCs possible, allowing exploration of various bioclinical as well as biodiagnostic applications.
ISSN:0141-8130
1879-0003
DOI:10.1016/j.ijbiomac.2016.11.076