Retinal Cryosectioning and Immunohistochemistry

Animals were euthanized at the end of experiment. Eyes were collected and processed for cryostat sections (10 µm) as described in our previous studies.⁴³ (link)^,⁴⁵ (link) Four sections/slide were collected in five series and stored at −80 °C. One slide from each series was stained with cresyl violet to assess the integrity of retinal lamination. Other slides were used for antibody staining, following previously described protocols, and were examined by regular light and confocal microscopy (Eclipse C1si; Nikon Instruments, Inc., Melville, NY). The following antibodies were used: CNTF (rabbit polyclonal, 1:500, Santa Cruz Biotechnology, Dallas, TX), Cone-arrestin (rabbit polyclonal, 1:1,000; Millipore, Burlington, MA), and protein kinase C-alpha (rabbit polyclonal, 1:5000; Sigma, St Louis, MO). Anti-rabbit secondary antibodies conjugated to Alexa Fluor-488 (Life Technologies, Carlsbad, CA) were used and sections were counterstained with 4′, 6-diamidino-2-phenylindole.

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Yang J.Y., Lu B., Feng Q., Alfaro J.S., Chen P.H., Loscalzo J., Wei W.B., Zhang Y.Y., Lu S.J, & Wang S. (2021). Retinal Protection by Sustained Nanoparticle Delivery of Oncostatin M and Ciliary Neurotrophic Factor Into Rodent Models of Retinal Degeneration. Translational Vision Science & Technology, 10(9), 6.

Publication 2021

Eclipse C1si Cone-arrestin Alexa Fluor-488

Alexa fluor 488 Animals Anti antibodies Antibodies Antibody Arrestin Cntf Cone Confocal microscopy Cresyl violet Eyes Light Protein kinase c alpha Rabbit Retinal

Corresponding Organization : Cedars-Sinai Medical Center

Other organizations : Beijing Tongren Hospital, Capital Medical University, Brigham and Women's Hospital, Harvard University

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Integrity of retinal lamination
Levels and localization of CNTF, cone-arrestin, and protein kinase C-alpha proteins

control variables

Retinal sections collected in five series and stored at -80 °C
One slide from each series stained with cresyl violet to assess retinal lamination
Other slides used for antibody staining following previously described protocols

controls

Positive control: Not specified
Negative control: Not specified

Annotations

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