Quantitative Fluorescent Imaging of RPE/Choroidal Flat Mounts

ISH of RPE/choroidal flat mounts was performed using the Multiplex Fluorescent Reagent Kit v2 (ACD) according to the protocol of Gross-Thebing et al. [26 (link)]. The following probes were used: Iba1 (channel C1 or C3), VEGF (channel C3), and PGF (channel C1). Images were taken with a Zeiss Imager M.2 including an ApoTome.2.
The colored pixel intensity in individual image areas of the laser spots was quantified using the Colored Pixel Counter tool for Fiji.

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Balser C., Wolf A., Herb M, & Langmann T. (2019). Co-inhibition of PGF and VEGF blocks their expression in mononuclear phagocytes and limits neovascularization and leakage in the murine retina. Journal of Neuroinflammation, 16, 26.

Publication 2019

Imager M.2 ApoTome.2

Choroidal Spots Vegf

Corresponding Organization : University of Cologne

Other organizations : University Hospital Cologne, Institute of Medical Microbiology and Hygiene

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Variable analysis

independent variables

Iba1 probe
VEGF probe
PGF probe

dependent variables

Colored pixel intensity in individual image areas of the laser spots

control variables

Multiplex Fluorescent Reagent Kit v2 (ACD) protocol
Zeiss Imager M.2 including an ApoTome.2 imaging system

controls

No positive or negative controls were explicitly mentioned in the provided information.

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