Monoclonal antibodies isolated by large-scale screening are suitable for labeling adult zebrafish (Danio rerio) tissues and cell structures.
The lack of a sufficient number of antibodies represents an obstacle in the research performed using the zebrafish (Danio rerio) as a model organism. On the other hand, high-throughput generation of antibodies, especially those suitable for immunohistochemistry, is not an established methodology. Here we present the results of an immunization experiment with a zebrafish tissue lysate that allowed for the isolation of hundreds of monoclonal antibodies suitable for labeling of a large variety of zebrafish tissue and cell structures. Some of them were further characterized in terms of detailed localization and age-dependent expression. In addition, the antigen recognized by one of them was first immunoprecipitated and then identified by mass spectrometry. Furthermore, immunofluorescence-competent recombinant antibodies were also isolated by panning large repertoire phage display libraries, in both single-chain (scFv) and single-domain (VHH) format. Such selection alternative is simpler to organize and could contribute to limit the costs of antibody screening and production.
Fondazione IFOM (Istituto FIRC di Oncologia Molecolare), Milan, Italy.
Gianluca
G
Deflorian
Mario
M
Cinquanta
Carlo
C
Beretta
Annunziata
A
Venuto
Cristina
C
Santoriello
Danila
D
Baldessari
Federica
F
Pezzimenti
Marisa
M
Aliprandi
Marina
M
Mione
Ario
A
de Marco
D000367:Age Factors; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D015153:Blotting, Western; D018604:Epitope Mapping; D006825:Hybridomas / Q000378:metabolism; D007114:Immunization; D007143:Immunoglobulin Heavy Chains / Q000235:genetics / Q000302:isolation & purification; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000302:isolation & purification; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D051379:Mice; D008807:Mice, Inbred BALB C; D008856:Microscopy, Fluorescence; D019151:Peptide Library; D011994:Recombinant Proteins / Q000302:isolation & purification; D015027:Zebrafish; D029961:Zebrafish Proteins / Q000032:analysis / Q000276:immunology*
D000911:Antibodies, Monoclonal; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D019151:Peptide Library; D011994:Recombinant Proteins; D029961:Zebrafish Proteins