Serum Proteome of Leprosy Patients Undergoing Erythema Nodosum Leprosum Reaction:  Regulation of Expression of the Isoforms of Haptoglobin

Abstract

Validated proteome profile allows better understanding of disease progression, subtype classification, susceptibility patterns, and disease prognosis. Leprosy is a spectral disease, with clinically, histologically, immunologically, and bacteriologically distinguishable subtypes. In addition, a significant fraction of patients undergo immune mediated reactions even after multidrug therapy (MDT). Erythema nodosum leprosum (ENL) is an immune complex mediated reactional condition in leprosy, characterized by a systemic inflammatory condition afflicting borderline lepromatous (BL) and lepromatous leprosy patients (LL). In this study, we have analyzed serum proteome of leprosy patients undergoing ENL reactions and compared it with that of healthy noncontact controls. Depletion of albumin and immunoglobulin G (IgG) was optimized using Aurum serum protein mini kit (Bio-Rad), and then two-dimensional gel electrophoresis (2-DE) of these serum samples was performed. Differentially expressed proteins were identified by MALDI-TOF and MALDI-TOF MS/MS mass spectrometry. Significant increase in one of the isoforms of alpha2 chain of haptoglobin was observed in ENL condition. In addition, haptoglobin phenotype was determined for healthy controls and leprosy patients. Hp 0-0 phenotype was detected in 21.4% of the ENL patients undergoing treatment, which on follow up examination showed typable phenotype, thus showing a condition of acquired anhaptoglobinemia. Since ENL still remains a threat to leprosy disease management, the above findings may provide new insights in understanding the development and progression of this inflammatory condition.

Preliminary proteomic analysis on the alterations in follicular fluid proteins from women undergoing natural cycles or controlled ovarian hyperstimulation

“…Vortexing and incubation were repeated three times. After these steps, the column was centrifuged at 10,000 g for 20 s. The unbound protein fraction was collected by washing the column again with 200 μL binding buffer, and the supernatant containing the albumin-and IgG-depleted FF sample was now ready for 2-DE [19].…”

“…Briefly, 450-μg aliquots of FF proteins were applied to immobilized pH 4-7 nonlinear gradient strips (18 cm, Amersharm Bioscience, Uppsala, Sweden), and the strips were rehydrated and focused as previously described [19]. The second-dimensional separation was performed in 12.5 % polyacrylamide gels.…”

Comparative proteomics and glycoproteomics of plasma proteins in Indian visceral leishmaniasis

“…Albumin and IgG were first depleted using the aurum serum protein mini kit (Bio-Rad, CA) [21]. Glycoproteins were purified from this depleted plasma using a multi lectin affinity chromatography (M-LAC) column by mixing 0.50 mL each of agarose-bound ConA, WGA and Jacalin lectin in a PD-10 disposable column (GE-Healthcare, USA).…”

“…The focused IPG-strips were equilibrated for 30 min each using equilibration buffer-I (6 M urea, 2 M thiourea, 0.375 M Tris–HCl, pH 8.8, 20% glycerol, 2% SDS, 0.005% bromophenol blue and 2% DTT) followed by buffer-II (2.5% iodoacetamide in place of DTT). The strips were then sealed on top of a gradient polyacrylamide gel with 0.5% agarose in electrophoresis buffer and separated using Bio-Rad gel apparatus [21,24]. …”

Dietary supplementation with selenomethylselenocysteine produces a differential proteomic response

“…Albumin depletion is a highly reproducible procedure that is frequently used in proteomics approaches to biomarkers discovery [23]. Protein concentration was determined according to the Bradford method using bovine serum albumin as protein standard.…”