Description of Research Expertise

o Practice of handling human subject:
 Blood collection from human vein (certified Phlebotomists).
 Experiment with normal, HIT, COVID-19, VITT and APS patient plasma sample.
o Practice of handling experimental laboratory animals (Rat, mice, guineapig, Rabbit, Toad):
 Handling mouse: Colonies maintenance, breeding and genotyping
 Dosage routes: Intraperitoneal, intramuscular, intravenous, intraarticular, intradermal, subcutaneous injection and gavage (force feeding) of drug/ minerals/ microRNA/ siRNA/adenovirus/nano-formulation.
 Anatomical measurements: Paw, knee, ankle diameter measurement from in rat and mice.
 Physiological parameters: Insulin and Glucose tolerance test in mice model. Echocardiography of mice, X-ray study, blood hematological parameters count.
 Surgery: Osteoporosis development, Cecal ligation puncture procedure (CLP model), Lung surgery model, Liver Surgery model, Cremaster model. Liver, lung surgery in mice model, along with injury, Cremaster muscle surgery along with injury of vessel in mice model.
 Model development: Osteoarthritis, Rheumatoid arthritis model development; Diabetes and Diabetic cardiomyophy model (Type I and Type II), Inflammation and nociception models in mice, Hyperimmunization model development (guineapig); LPS induced Sepsis model, Thrombosis model, In vitro COVID-19 and Vaccine induced thrombotic thrombocytopenia (VITT) model.
 Fluid collection: Synovial fluid collection from joint and its analysis through ELISA, Blood collection (hepatic vein, vena cava, retro-orbital, cardiac, tail vein, jugular vein, marginal ear vein (rabbit), iliac vein)
 Toxicity studies: in animal (Hemolysis, hemorrhagic study, acute toxicity study).
 Pharmacological Studies: Drug dose response in isolated guineapig heart auricle, Drug dose response in isolated rat phrenic nerve, Drug dose response in toad heart.
 Biochemical analysis: Physiological markers analysis from blood, urine from rat and mice model. Oxidative stress molecule analysis from rat and mice blood. Free DNA measurement from mice plasma, free DNA assessment from mice plasma.
o Cell culture techniques:
 Isolation, culture and treatment of primary cell culture (synoviocyte culture both in human and rat, primary neonatal rat cardiomyocyte, primary adult mice and rat cardiomyocyte, Lymphocyte isolation from spleen, heart and blood), B, T cell and mice and human monocyte isolation and culture using isolation kit.
 Handling and treatment of HIG-82 (synoviocytes) cell line, H9C2 and C2C12 (cardiomyocyte) cell line, A20 (B lymphoma) cell line, HUVEC primary cell line, mouse aortic endothelial cells line, porcine WT kidney endothelial cells, porcine aortic endothelial cells.
 High and low glucose treatment and analysis in cardiomyocyte (both primary and stable cell line).
 Hybridoma cells for antibody production.
 MTT assay for cell apoptosis determination.

o Antibody production from hybridoma cells:
 Antibody production, purification from plasma and hybridoma sup.
 Antibody labelling.
 IgG purification, digestion and Fc-modification, preparation of F(ab’)2 fragment.
o B cell Biology:
 Evaluation of B cell expression in heart section using immunohistochemistry, immunofluorescence staining.
 Evaluation of B cell expression on heart tissue using western blot and immunoprecipitation technique
 Evaluation of B cell expression using flow cytometry.
 Evaluation of B cell expression and B cell homing from circulation to heart using B cell specific gene therapy.
o Bioflux microfluidics:
 Microfluidics model and drug interaction after photochemical injury in cell culture using human, mouse, porcine endothelial cells for in vitro thrombosis model.
o COVID and VITT experiment:
 Handling COVID and VITT plasma and run them in microfluidics system for thrombosis experiment.
o Molecular Biology:
 Isolation of total RNA and cDNA synthesis.
 RT-PCR, Real Time PCR.
 Agarose or polyacrylamide gel electrophoresis
 Primer design for RT-PCR and qPCR
 Microarray analysis.
o Protein Biochemistry:
 Protein estimation and preparation.
 Protein expression and interaction: Gel electrophoresis (PAGE and SDS), Western blot, immuno-precipitation and co-immuno- precipitation.
 Sandwich ELISA.
 HPLC, UV Scan, CD spectra analysis, Fluorescence study, X-RD analysis, FTIR analysis, EDAX, MALDI-TOF, LC-MS/MS, Dialysis of protein sample for salting in and out.
o Carbohydrate analysis:
 Thin Layer Chromatography, Paper chromatography, acid hydrolysis for carbohydrate separation and identification.
o Tissue handling and staining:
 Tissue dehydration and block preparation and sectioning
 Hematoxylin and eosin staining; immunohistochemical (single and double color staining), immunofluorescence staining and imaging of tissue sections.

o Cell staining:
 Immunocytochemistry and immunofluorescence (double color) staining and imaging.
 Confocal imaging of live cells using trackers for colocalization.
 Synoviocyte apoptosis measurement by annexin V-FITC through fluorescent microscopy.
 Immunofluorescence study for apoptosis detection.
o Flow cytometry:
 Antibody expression detection using 4 colors and 8 colors together, Cell apoptosis study.
o Microscopy:
 Bright light microscopy, fluorescence microscopy, scanning electron microscopy, Live cell imaging using EVOS, confocal microscopy both with cells and in live mice under anesthesia and in live cell and animal.
o Nanoformulation and Nanotechnology:
 Gold nanoparticle preparation and tagging with protein, tagging with fish oil lipid and peptide and encapsulation within liposome to target specific to arthritis in both animal model and synoviocyte cell cultures.
 Liposome preparation and characterisation
 Characterization of nanoparticle- DLS, Zeta, TEM and SEM imaging, EDAX, Contact angle to analyse hydrophobicity or hydrophilicity of the particle.
o Bacteriology:
 Colony forming unit from mouse blood and organs.
o Software Specific Skills:
 Origin lab Pro.
 GraphPad Prism.
 Bioinformatics tools: BLAST-protein and nucleotide.
 Microsoft Office, Excel, Power Point.
 Adobe Photoshop, Image J (western blot, immunocytochemistry, immunofluorescence, confocal analaysis)
o Hands on training
• Human Cell Culture, FPLC (Chromatography), Gel Filtration, X- Ray Crystallography by hanging drop vapour diffusion method from Goa BITS Pillani, Goa, India.