IHC was performed at Indivumed. Tumor samples were fixed in 4% neutral buffer formaldehyde solution (10% formalin) and transferred to 70% ethanol at Incyte Corporation. Subsequently, the formalin-fixed samples were embedded in paraffin at Indivumed according to Indivumed's standard operating procedure. IHC was implemented on the Discovery XT staining platform (Roche Diagnostics/Ventana Medical Systems), using the rabbit monoclonal anti–PD-L1 antibody clone SP142 (Spring BioScience/Ventana). Formalin-fixed, paraffin-embedded tissue samples were sliced into 3- to 5-µm sections and mounted on Superfrost Ultra Plus glass slides (Carl Roth). Sections were deparaffinized within the staining instrument and immunostained using the Discovery ChromoMap DAB Kit (Roche Diagnostics). After staining, the slides were manually washed using hot tap water supplemented with detergent, followed by tap water only and distilled water in a final step. For dehydration, the slides were transferred to an ascending ethanol series (2 × 80%, 2 × 96%, 2 × abs. EtOH; 1 minutes each). After dehydration, the slides were transferred to xylene (2 × 1 minutes) and automatically cover-slipped in Pertex mounting medium (International Medical Products). Scans were generated with the Axio Scan Z1 automated slide scanner (Zeiss) using ZEN 2 (blue edition) slide scan software (Zeiss).
Zen 2 blue edition slidescan software
Manufactured by Zeiss
ZEN 2 (blue edition) is a slidescan software developed by Zeiss. It provides a core functionality for digitizing microscope slides.
Automatically generated - may contain errors
Lab products found in correlation
Triton x 100, by Merck Group (2 mentions)
Axio scan z1 automated slide scanner, by Zeiss (2 mentions)
Polysine microscopic glass slides, by Thermo Fisher Scientific (2 mentions)
Donkey serum, by GeneTex (2 mentions)
Axio scan z1, by Zeiss (1 mentions)
Discovery chromomap dab kit, by Roche (1 mentions)
3 protocols using zen 2 blue edition slidescan software
1
PD-L1 Immunohistochemistry Protocol
2
Anatomical Characterization of Mouse Brain
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Adult mice were deeply anesthetized with 2% avertin and perfused with 0.1 M PBS followed by 4% paraformaldehyde. Brains were postfixed in 4% paraformaldehyde at 4°C for 24 hours and 30% sucrose at 4°C for 48 hours. Frozen brains were then cut into 30 μm coronal cryosections. Sections were blocked in 0.1 M PBS containing 0.3% Triton X-100 (Sigma) and 2% Donkey Serum (GeneTex) for 30 min at room temperature. Primary antibody was applied at the appropriate dilution and incubated overnight at 4°C. Incubated sections were washed three times with 0.1 M PBS and incubated in secondary antibody for two hours. After three rinses in 0.1 M PBS and DAPI staining at 1:1000 (Pierce), the sections were mounted on polysine microscopic glass slides (Thermo Scientific). Images were scanned with the Axio Scan Z1 automated slide scanner (Zeiss) using ZEN 2 (blue edition) slidescan software (Zeiss).
3
Anatomical Characterization of Mouse Brain
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Adult mice were deeply anesthetized with 2% avertin and perfused with 0.1 M PBS followed by 4% paraformaldehyde. Brains were postfixed in 4% paraformaldehyde at 4°C for 24 hours and 30% sucrose at 4°C for 48 hours. Frozen brains were then cut into 30 μm coronal cryosections. Sections were blocked in 0.1 M PBS containing 0.3% Triton X-100 (Sigma) and 2% Donkey Serum (GeneTex) for 30 min at room temperature. Primary antibody was applied at the appropriate dilution and incubated overnight at 4°C. Incubated sections were washed three times with 0.1 M PBS and incubated in secondary antibody for two hours. After three rinses in 0.1 M PBS and DAPI staining at 1:1000 (Pierce), the sections were mounted on polysine microscopic glass slides (Thermo Scientific). Images were scanned with the Axio Scan Z1 automated slide scanner (Zeiss) using ZEN 2 (blue edition) slidescan software (Zeiss).
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