title: A novel approach for the chromatographic purification and peptide mass fingerprinting of urinary free light chains.
abstract: 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.
authors.affiliation: Laboratory of Native Proteins, Research and Development Division, Yashraj Biotechnology Ltd., Navi Mumbai, 400705 Maharashtra, India.
authors.forename: Bhupesh C
authors.initials: BC
authors.lastname: Mali
authors.affiliation: Laboratory of Native Proteins, Research and Development Division, Yashraj Biotechnology Ltd., Navi Mumbai, 400705 Maharashtra, India. Electronic address: sham83badgujar@gmail.com.
authors.forename: Shamkant B
authors.initials: SB
authors.lastname: Badgujar
authors.affiliation: Laboratory of Native Proteins, Research and Development Division, Yashraj Biotechnology Ltd., Navi Mumbai, 400705 Maharashtra, India.
authors.forename: Kunal K
authors.initials: KK
authors.lastname: Shukla
authors.affiliation: Laboratory of Native Proteins, Research and Development Division, Yashraj Biotechnology Ltd., Navi Mumbai, 400705 Maharashtra, India.
authors.forename: Paresh B
authors.initials: PB
authors.lastname: Bhanushali
mesh_terms: D000595:Amino Acid Sequence; D002845:Chromatography / Q000379:methods*; D006801:Humans; D007145:Immunoglobulin kappa-Chains / Q000737:chemistry / Q000302:isolation & purification* / Q000652:urine*; D007146:Immunoglobulin lambda-Chains / Q000737:chemistry / Q000302:isolation & purification* / Q000652:urine*; D008506:Medical Waste; D010449:Peptide Mapping / Q000379:methods*
chemical_list: D007145:Immunoglobulin kappa-Chains; D007146:Immunoglobulin lambda-Chains; D008506:Medical Waste
keywords: Bence Jones proteins; Free light chains; Immunoblotting; Proteomics; Urine