Development and characterisation of a monoclonal antibody family against aquaporin 1 (AQP1) and aquaporin 4 (AQP4).
Recent studies have discovered the existence of water-channel molecules, the so called aquaporins (AQP) presumably involved in active, ATP dependent water transport between the intracellular and extracellular compartments. Both genetic and protein sequences and structures of the AQPs are known and crystallographic analyses of some members of the AQP family have been performed. Specific antibodies are required to examine their histological locations and analyse their roles in physiological and pathological pathways of water transportation and osmotic regulation. Until recently some polyclonal antibodies have been developed against certain members of the AQP family. However, to date highly specific monoclonal antibodies against aquaporins do not exist. We have developed a monoclonal antibody family against the aquaporin 1 (AQP1) and aquaporin 4 (AQP4) molecules. Well-conserved epitop sequences of AQP1 and AQP4 proteins were selected by computer analysis and their synthetic peptide fragments were used as the antigens of immunisation and the following screening. Antibodies were characterised by immunoserological methods (ELISA, dot-blot and immunoblot), flow cytometry and immunohistochemistry of formaldehyde-fixed and paraffin-embedded tissue samples. RT-PCR tests controlled the specificity of the immune reactions. Our monoclonal antibodies recognised AQP1 and AQP4 in all the techniques mentioned above and apparently they are useful both in various quantitative and qualitative measurements of the expressions of AQP1 and AQP4 in several species (human, rat, mouse, invertebrates, even plants). According to preliminary immunohistochemical studies our monoclonal anti-AQP1 and anti-AQP4 antibodies are appropriate tools of patho-morphological examinations on routine formol-paraffin tissue samples.
Nagy
Gergely
G
Department of Immunology and Biotechnology, University of Pécs, Faculty of Medicine, Pécs, H-7643, Hungary. peter.nemeth@aok.pte.hu
Szekeres
György
G
Kvell
Krisztián
K
Berki
Tímea
T
Németh
Péter
P
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000302:isolation & purification; D051398:Aquaporin 1; D051401:Aquaporin 4; D020346:Aquaporins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000276:immunology / Q000473:pathology; D017931:DNA Primers / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005434:Flow Cytometry; D005753:Gastric Mucosa / Q000378:metabolism; D006825:Hybridomas; D007114:Immunization; D015151:Immunoblotting; D007124:Immunoenzyme Techniques; D007668:Kidney / Q000378:metabolism; D008113:Liver Neoplasms / Q000276:immunology / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D016612:Paraffin Embedding; D010446:Peptide Fragments / Q000276:immunology; D012313:RNA / Q000302:isolation & purification / Q000378:metabolism; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured / Q000473:pathology
D000911:Antibodies, Monoclonal; C496021:Aqp1 protein, mouse; C496022:Aqp1 protein, rat; C496031:Aqp4 protein, mouse; C496032:Aqp4 protein, rat; D051401:Aquaporin 4; D020346:Aquaporins; D017931:DNA Primers; D010446:Peptide Fragments; D051398:Aquaporin 1; D012313:RNA