An Immunohistochemical Analysis to Validate the Rationale behind the Enhanced Immunogenicity of D-Ribosylated Low Density Lipo-Protein

Abstract

Advanced glycation end products (AGEs) are thought to contribute to the abnormal lipoprotein profiles and increased risk of cardiovascular disease in patients with diabetes and renal failure. D-ribose is one of the naturally occurring pentose monosaccharide present in all living cells and is a key component of numerous biomolecules involved in many important metabolic pathways. Formation of D-ribose derived glycated low density lipoprotein (LDL) has been previously demonstrated but no studies have been performed to assess the immune complex deposition in the kidney of rabbits immunized with glycated LDL. In this study, LDL was glycated with D-ribose, and it was further used as an immunogen for immunizing NZW female rabbits. The results showed that female rabbits immunized with D-ribose modified LDL induced antibodies as detected by direct binding and competitive ELISA. The modified LDL was found to be highly immunogenic eliciting high titer immunogen-specific antibodies, while the native forms were moderately immunogenic. The induced antibodies from modified LDL exhibited wide range of heterogeneity in recognizing various proteins and amino acids conformers. Furthermore, our histopathological results illustrated the deposits of immune complex in glomerular basement membrane in rabbits immunized with D-ribose-LDL.

Biochemical and immunological parameters as indicators of osteoarthritis subjects: role of OH-collagen in auto-antibodies generation

“…To analyze the role of OH-collagen II in eliciting immune response in OA patients, the presence of auto-antibodies in arthritis subjects against OH-collagen II was probed by direct binding ELISA (Shahab et al, 2013[ 29 ]; Ahmad et al, 2011[ 5 ]; Moinuddin et al, 2014[ 23 ]; Akhter et al, 2014[ 11 ][ 13 ]). Out of 100 arthritic sera (50 male and female each), 30 male patient samples (60 %) showed higher binding with the OH-collagen II as compared to the native form, however, 40 female subjects (80 %) showed enhanced binding.…”

Recent advances in detection of AGEs: Immunochemical, bioanalytical and biochemical approaches

“…Presences of antibodies against Amadori‐rich glycated poly‐ l ‐lysine have been shown in sera of diabetic patients . Autoantibodies against glycated form of IgG, BSA, LDL and HSA have also been observed and detection of these autoantibodies can aid in the detection of their respective glycated proteins which otherwise is ambiguous due to the lack of universally accepted techniques . Therefore, it is important to focus on their detection as it would greatly assist in early diagnosis of glycation‐associated diseases.…”

Detection of Circulating Auto-Antibodies Against Ribosylated-LDL in Diabetes Patients

“…Direct binding ELISA was performed on polystyrene plates with slight adjustment . Polystyrene immunoplates were covered with 100 μl of the native or glycated LDL (10 μg/ml) in 0.05 M carbonate–bicarbonate buffer with pH 9.6.…”

“…Immunoglobulin G (IgG) was affinity purified from normal human sera (NHS) and T1DM, T2DM patient's sera on a protein A‐Sepharose column . Serum (0.5 ml) diluted with equivalent volume of PBS (pH 7.4), was applied to the column pre‐equilibrated with above buffer.…”