Intracelluar infections- finding target molecules on the surface of pathogen and infected cells using monoclonal antibodies. Infectious diseases, malaria and tuberculosis in particular, remain the leading cause of human mortality. Central to our understanding the various mechanisms involved in the pathogenesis of the disease are identification of the pathogen and infected cell associated molecules. The objectives of our ongoing efforts are two folds: i) to identify the neo-antigenic determinants on pathogen and infected cell- surface and, ii) to characterize the identified molecules biochemically and functionally, using monoclonal antibodies (MAbs) generated by employing novel immunization strategies. Taking malaria infected erythrocytes, leishmania and mycobacteria- infected macrophages as model systems, for the first time we showed that the antibodies, generated by immunization with the parasite-infected homologous cell membranes, react with new antigenic determinants on pathogen and infected cell surface both in vitro and in vivo. Also, such immunization strategy was found useful in diverting the immune response towards functionally important “minor” antigenic determinants. In malaria infected erythrocytes we also established the rationale that the specific determinants on the infected cell surface can be exploited for selective elimination of the diseased cells. After generating panels of monoclonal antibodies, we have identified few target molecules on above mentioned pathogens and respective infected cell surface. The focus of the ongoing studies are to characterize these molecules using immunological, bioinformatics and molecular tools. The concepts of the antibody specificity and cross reactivity against small molecules. Studies on the cross-reactivity and specificity of antibodies are important not only serologically but central to delineate the epitopic and functional regions on the antigen. Our understanding towards the serological reactions have evolved over the decades since the classic work of Landsteiner. Unlike macromolecules, which elicit a multiplicity of antibody subpopulations towards the different regions of the macromolecule surface, small molecules (haptens) usually elicit antibodies when bound to appropriate carriers which are complementary to one or a few functional groups. Based on our recent interesting observations on the antibody specificities against small molecules, in collaborative efforts, our long term goal is to identify functional group(s) in smaller molecules using antibodies. These studies are not only of fundamental in nature but also would aid in designing vaccines and specific diagnostic kits against small molecules. Applied studies. Work in this direction involves the development of monoclonal antibodies having potential in diagnostics and therapeutics. Significant recognition: Awards, fellowships, international funding of distinction, technologies transferred/licensed etc.: Technology Development : Licensed Technology, entitled: “Muc 4 Antibodies for Research Use” Batra, S.K.; Brand, R.E.; Ringel, J.; Faulmann, G.; Lohr, M. and Varshney, G.C. US Patent No. 7052859 B2 (2006) Muc 4 Antibody for research use (8G7) is being marketed by Abcam, Biocare Medical and Santa Cruz. Awards : Research Innovation Award 2007 from UneMed, USA (UneMed Corporation and University of Nebraska Medical Center,) for Licensed Technology, entitled: “Muc 4 Antibodies for Research Use” DBT, Govt. of India, Overseas Research Associateship (Aug., 2000-Jan., 2001) Selected list of Publications and Patents: Selected research publications: Verma R, Tiwari A, Kaur S, Varshney GC and Raghava GPS (2008). Identification of Proteins Secreted by Malaria Parasite into Erythrocyte using SVM and PSSM profiles. BMC Bioinformatics, 9, 201-211 Moniaux N, Chaturvedi P, Varshney GC, Meza JL, Rodriguez-Sierra JF, Aubert JP and Batra SK (2007). Human MUC4 mucin induces ultra-structural changes and tumorigenicity in pancreatic cancer cells British J. Cancer 97(3):345-57. Kaur J, Singh KV, Boro R, Thampi KR, Raje M, Varshney GC and Suri CR, (2007). Immunochromatographic dipstick assay format using gold nanoparticles labeled protein-hapten conjugate for the detection of atrazine. Environmental Science & Technology , 41, 5028-5036 Andrianifahanana M, Singh A. P, Nemos C, Ponnusamy M. P, Moniaux N, Mehta P. P, Varshney G.C and Batra S. K. (2007). IFN-×¥ -induced expression of MUC4 in pancreatic cancer cells is mediated by STAT-1 upregulation: a novel mechanism for IFN- ×¥ response. Oncogene, 26 (51): 7251- 7261. Pawaria S, Rajamohan G , Gambhir V, Lama A, Varshney G.C and Dikshit K.L (2007). Intracellular growth and survival of Salmonella enterica serovar Typhimurium carrying truncated hemoglobins of Mycobacterium tuberculosis. Microbial Pathogenesis, 24, 175-183. Singh M.K., Srivastava S., Raghava G. P. S. and Varshney G.C. (2006). HaptenDB : Collection and Compilation of Small antigenic Molecules. Bioinformatics, 22 (2), 253-256. Singh M.K., Srivastava S., Raghava G. P. S. and Varshney G.C. (2004). Hapten DB . Nucleic Acid Research (online). Suri, C.R., Raje, M and Varshney, G.C. (2002). Immunosensor for pesticide analysis: antibody production and sensor development . Critical Reviews in Biotechnology. 22 (1), 15-32. Sarin, J., Aggarwal, S., Chaba, R., Varshney, G.C. and Chakraborti, P.K. (2001). -subunit of phosphate specific transporter from Mycobacterium tuberculosis is a thermostable ATPase. J. Biol. Chem. 276, 44590-44597. Jain, M., Karan, D., Batra, S.K. and Varshney, G.C. (2001). Mucins in protozoan parasites. Frontiers in Biosciences, 6, d1276-1283. Majumdar, S., Kaur, H., Vohra, H. and Varshney, G.C. (2000). Membrane surface of Mycobacterium microti infected macrophages antigenically differs from that of uninfected macrophages. FEMS Immunol. Med. Microbiol. 28, 71-77. Goel, A., Vohra, H. and Varshney, G.C. (1999). Strain-specific recognition of live Leishmania donovani promastigotes by homologous antiserum raised against a crude membrane fraction of infected macrophages. Parasitol . Res. 85, 19-24. Choudhury, A., Goel, A., Raje, M., Vohra, H and Varshney, G.C. (1997). Recognition of the parasite infected cell surface determinants by homologous antiserum raised against infected cell membranes. Parasitol. Res. 83, 746-754.. Patents: Batra, S.K.; Brand, R.E.; Ringel, J.; Faulmann, G.; Lohr, M. and Varshney, G.C. MUCIN 4 expression as a marker for pancreatic cancer. US Patent No. 7052859 B2 (2006). Batra, S.K.; Brand, R.E.; Ringel, J.; Faulmann, G.; Lohr, M. and Varshney, G.C. Specific mucin expression as a marker for pancreatic cancer. US Patent No. 6,576,423 2003). Suri, C.R.; Raje, M. and Varshney, G.C. An efficient and improved method for screening of analytes by microporous filtration based dot immunoassay kit. European Patent 1579211B1 (2007) and Australian Patent 2002348763B2 (2007). Gupta, C.M., Owais, M. and Grish C. Varshney. A process for the preparation of drug encapsulated target specific immunoliposomes for the treatment of drug resistant diseases. Indian Patent No. 182550 (1999). Grish C. Varshney, Mohan Singh & Amit Choudhry. A process for the preparation of therapeutically useful novel monospecific proteins active against parasite infected cells. Indian Patent No. 182589 (1999). Book Chapter: Srivastava.S.,Singh.M.K.,Raghava G.P.S and Varshney.G.C.(2007). Searching haptens, carrier proteins and anti-hapten antibodies. In: Flower, DR. (Ed.). Immunoinformatics.:predicting immunogenicity in silico. Series: Methods in Molecular Biology, Humana Press, 409, 125-139.

Cell biology and immunology, hybridoma technology and antigen-antibody interactions