Retinas were dissected, and wholemounts were prepared by making four relieving cuts, one in each quadrant, to permit the retina to lay flat. Retinal wholemounts or transverse sections were then stained with isolectin GS-IB4 conjugated to Alexa-647 (I32450, ThermoFisher Scientific), diluted 1:100 to label blood vessels and microglial cells and immunostained using a combination of antibodies to the following antigens:; Brn3b (sc6026 Santa Cruz), diluted 1:250 to label retinal ganglion cells; Desmin (sc7559, Santa Cruz), diluted 1:500, and NG2 (gift from W. Stallcup lab in Sanford Burnham Prebys) [31 (link)], diluted 1:1000 to label pericytes; Iba1 (016-26461, Wako), diluted 1:1000 to label microglia; GFAP conjugated to Cy3 (C9205, Sigma), diluted 1:400 to label astrocytes and reactive Müller glia, and Collagen IV (AB769 Millipore), diluted 1:500 to label blood vessels. The secondary antibodies were raised in donkey, conjugated to IgG-Alexa488, IgG-Alexa594 or IgG-Alexa647 (ThermoFisher Scientific), diluted 1:200. All immunostaining steps included incubation with 5% donkey serum for 24 hours, primary antibodies for 3 days, and secondary antibodies for 24 hours, all at 4˚C with gentle agitation. The antibodies were diluted in 1% triton-PBS.
Igg alexa594
Manufactured by Thermo Fisher Scientific
IgG-Alexa594 is a fluorescently labeled antibody that can be used for various laboratory applications. It consists of an IgG antibody conjugated with the Alexa Fluor 594 fluorescent dye. The core function of this product is to provide a means of detecting and visualizing target molecules or structures through fluorescence microscopy or other fluorescence-based techniques.
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Lab products found in correlation
2 protocols using igg alexa594
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Retinal Wholemount Immunostaining
2
Retinal Wholemount Immunostaining
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Retinas were dissected, and wholemounts were prepared by making four relieving cuts, one in each quadrant, to permit the retina to lay flat. Retinal wholemounts or transverse sections were then stained with isolectin GS-IB4 conjugated to Alexa-647 (I32450, ThermoFisher Scientific), diluted 1:100 to label blood vessels and microglial cells and immunostained using a combination of antibodies to the following antigens:; Brn3b (sc6026 Santa Cruz), diluted 1:250 to label retinal ganglion cells; Desmin (sc7559, Santa Cruz), diluted 1:500, and NG2 (gift from W. Stallcup lab in Sanford Burnham Prebys) [31 (link)], diluted 1:1000 to label pericytes; Iba1 (016-26461, Wako), diluted 1:1000 to label microglia; GFAP conjugated to Cy3 (C9205, Sigma), diluted 1:400 to label astrocytes and reactive Müller glia, and Collagen IV (AB769 Millipore), diluted 1:500 to label blood vessels. The secondary antibodies were raised in donkey, conjugated to IgG-Alexa488, IgG-Alexa594 or IgG-Alexa647 (ThermoFisher Scientific), diluted 1:200. All immunostaining steps included incubation with 5% donkey serum for 24 hours, primary antibodies for 3 days, and secondary antibodies for 24 hours, all at 4˚C with gentle agitation. The antibodies were diluted in 1% triton-PBS.
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