Dissection and labelling of retinas was performed as previously described27 (link). Briefly, retinas were fixed for 2 h on ice in 4% paraformaldehyde, incubated in 1% BSA and 0.3% Triton X-100, washed two times in Pblec (1% Triton X-100, 1 mM CaCl2, 1 mM MgCl2, and 1 mM MnCl2 PBS (pH 6.8)), and incubated overnight with isolectin-B4 (1:50) and antibodies diluted in Pblec. Images were acquired and processed using a Leica TCS SP5 II microscope, LAS Montage Imaging software (Leica) and the IMARIS Digital Imaging software (Biplane). Biotinylated Griffonia simplicifolia lectin I (IB4), (B-1205) was purchased from Vector Laboratories.
AntibodiesDissection Griffonia simplicifolia lectin i Imaris Isolectin Mgcl2 Microscope Mncl2 Paraformaldehyde Retinas Triton x 100 Vector
Corresponding Organization :
Other organizations :
Sanquin, University of Amsterdam, UConn Health, William Harvey Research Institute, Queen Mary University of London, Netherlands Institute for Neuroscience, Ludwig-Maximilians-Universität München
Incubation with isolectin-B4 (1:50) and antibodies diluted in Pblec
dependent variables
Retina morphology and structure
control variables
Retina fixation for 2 h on ice in 4% paraformaldehyde
Washing in Pblec (1% Triton X-100, 1 mM CaCl2, 1 mM MgCl2, and 1 mM MnCl2 PBS (pH 6.8))
positive controls
Biotinylated Griffonia simplicifolia lectin I (IB4), (B-1205) from Vector Laboratories
negative controls
Not explicitly mentioned
Annotations
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