Tips and tricks about immunolocalization

On 21st of April Dr. Anna Gustavsson from Umeå Plant Science Centre presented at Agrisera a technical seminar about immunolocalization methods tips and tricks. Anna works daily with immunolocalization of various proteins in Arabidopsis thaliana roots using confocal laser scanning microsope. Her experience covers not only plant but also mammalian tissues. She shared with Agrisera staff many insights into immunolocalization procedure.
Thank you very much for this valuable technical seminar.

The image below is showing why pre-immune serum screening is of crucial importantce before starting custom antibody production. Serum from some animals may not be suitable at all due to increased background levels already before immunization is started.

Prescreening for suitable rabbits for antibody production for immunofluorescence of plants. Preserum of 10 different rabbits were screened by immunofluorescence of 5-day old roots of Arabidopsis thaliana ecotype Col-0 (WT). The preserum were diluted to 1:500 and 1:2500 and the secondary antibody, CY5-coupled donkey anti-rabbbit (Jackson ImmunoResearch) was diluted 1:300. Rabbit anti-KNOLLE at a dilution of 1:4000 (Lauber MH et al. 1997. J Cell Biol 139: 1485–1493) was used as a positive control and incubation with secondary antibody without primary was used as a negative control.

Courtesy of Dr. Anna Gustavsson and Prof. Markus Grebe, Umeå Plant Science Centre, Sweden