Development of a ligand blot assay using biotinylated live cells.
Adhesive interactions between cells are critical to a variety of processes, including host-pathogen relationships. The authors have developed a new technique for the observation of binding interactions in which molecules obtained from excised tissues are resolved by gel electrophoresis and transferred to a membrane. Biotinylated live cells are then kept in contact with that membrane, and their interactions with proteins of interest are detected by peroxidase-labeled streptavidin, followed by a biotin-streptavidin detection system. The adhesion proteins can eventually be identified by cutting the relevant band(s) and performing mass spectrometry or other amino acid-sequencing methods. The technique described here allows for the identification of both known and novel adhesion molecules capable of binding to live cells, among a complex mixture and without previous isolation or purification. This is especially important for the analysis of host-parasite interactions and may be extended to other types of cell-cell interactions.
De Almeida Dias
Felipe
F
Instituto de Microbiologia Prof. Paulo de Góes, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.
Souza Dos Santos
André Luis
AL
Ferreira-Pereira
Antônio
A
Romeiro
Alexandre
A
Teixeira Zimmermann
Luciana
L
Cunha do Nascimento
Michelle Tanny
MT
Correa Atella
Georgia
G
Saraiva
Elvira Maria
EM
Linden
Rafael
R
Hampshire Lopes
Angela
A
D000818:Animals; D001710:Biotin / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006790:Host-Parasite Interactions; D007313:Insecta; D008024:Ligands
D008024:Ligands; D001710:Biotin