The 3μm thick paraffin-embedded kidney sections were dewaxed, and then dehydrated using graded concentrations of alcohol. To inhibit endogenous peroxidase, the sections were incubated with 3% H2O2. The sections were microwaved in citric acid for 15 min, and then treated with goat serum for 15 min at room temperature. Afterwards, the sections were incubated in blocking solution with primary antibody at 4°C overnight. After washing with PBS 3 times, the secondary antibody was added and immunostaining was performed using a DAB horseradish peroxidase color development kit (Beyotime, China), and then sections were counterstained with hematoxylin and made transparent with xylene. Finally, sections were observed with the PD37 type microscope (Olympus,Japan). Under 400 × magnification, pictures were taken in 5 random fields. Primary antibodies were used at the following dilutions: IL-1β diluted 1:100 (WL02257, Wanlei, Shenyang, China); IL-18 diluted 1:100 (WL01127, Wanlei, Shenyang, China).
Pd37 type microscope
Manufactured by Olympus
Sourced in Japan
The PD37 type microscope is a compact and versatile laboratory instrument designed for various applications. It features a binocular observation head, allowing users to view samples with both eyes. The microscope is equipped with adjustable interpupillary distance and diopter settings to accommodate individual user needs. The PD37 utilizes a halogen illumination system to provide bright and even illumination of the sample. It is capable of operating at multiple magnification levels to suit different imaging requirements.
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Lab products found in correlation
Dab horseradish peroxidase color development kit, by Beyotime (1 mentions)
Il 18, by Wanlei (1 mentions)
Il 1β, by Wanlei (1 mentions)
Anti ampk, by Abcam (1 mentions)
Dab horseradish peroxidase color development kit, by Boster Bio (1 mentions)
Anti p62, by Boster Bio (1 mentions)
Anti beclin1, by Boster Bio (1 mentions)
Anti mtor, by Abcam (1 mentions)
2 protocols using pd37 type microscope
1
Immunostaining of Kidney Sections for IL-1β and IL-18
2
Immunohistochemical Analysis of Autophagy Markers
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After the paraffin-embedded sections were dewaxed, we transferred the slides to 100% alcohol, 95% alcohol, 75% alcohol, and ddH 2 O 2 min twice respectively. After antigen retrieval, put them in hydrogen peroxide (3%) for 10 min to eliminate endogenous peroxidase activity. After washing with PBS three times, the sections of different groups were incubated with anti-AMPK (1:100, abcam, Cambridge, UK), anti-mTOR (1:400, abcam, Cambridge, UK), anti-ULK1 (1:300, Boster, Wuhan, Hubei, China), anti-Beclin-1 (1:200, Boster, Wuhan, Hubei, China), anti-p62 (1:200, Boster, Wuhan, Hubei, China), anti-LC3-II/LC3-I (1:100, Boster, Wuhan, Hubei, China) at 37 for 1 h. After washing with PBS three times, the secondary antibody (Boster) was added and immunostaining was performed using a DAB horseradish peroxidase color development kit (Boster, Wuhan, Hubei, China), and then, sections were counterstained with hematoxylin and made transparent with xylene.
Finally, sections were observed with the PD37 type microscope (Olympus, Japan). Under 20× magnification, pictures were taken in five random fields. The catalog numbers: anti-ULK1 (A00584-1), anti-Beclin-1 (PB9076), anti-LC3-II/LC3-I (BM4827), anti-p62 (PB0458), anti-AMPK (ab32047), and anti-mTOR (ab32028).
Finally, sections were observed with the PD37 type microscope (Olympus, Japan). Under 20× magnification, pictures were taken in five random fields. The catalog numbers: anti-ULK1 (A00584-1), anti-Beclin-1 (PB9076), anti-LC3-II/LC3-I (BM4827), anti-p62 (PB0458), anti-AMPK (ab32047), and anti-mTOR (ab32028).
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