Immunofluorescence analysis was performed according to protocols. Cells were implanted in 24-well dishes and fixed by 4% paraformaldehyde 24 h later. Fixed cells were stained with autophagy-related proteins (Cell Signaling Technology, USA), Rage (Cell Signaling Technology), followed by FITC-conjugated anti-mouse IgG and Cy3-conjugated anti-rabbit IgG (Abcam). Representative images were detected by fluorescent microscopy (Leica, German) and data were processed via ImagePro Plus.
Image pro plus
Manufactured by Leica camera
Sourced in Germany
Image-Pro Plus is a comprehensive image analysis software developed by Leica. It provides a range of tools for measuring, counting, and analyzing digital images. The software is designed to work with a variety of image formats and can be used for various applications, such as materials science, life sciences, and industrial inspection.
Automatically generated - may contain errors
Lab products found in correlation
Fluorescent microscopy, by Leica camera (4 mentions)
Cy3 conjugated anti rabbit igg, by Abcam (3 mentions)
Fitc conjugated anti mouse igg, by Abcam (2 mentions)
Autophagy related proteins, by Cell Signaling Technology (1 mentions)
Goat anti rabbit igg antibody, by Thermo Fisher Scientific (1 mentions)
Fluorescence microscope, by Leica camera (1 mentions)
Fluorescence microscopy, by Leica camera (1 mentions)
Fitc conjugated anti rabbit igg, by Cell Signaling Technology (1 mentions)
4 6 diamidino 2 phenylindole dapi, by Abcam (1 mentions)
Dfc300fx, by Leica Microsystems (1 mentions)
Aleuria aurantia lectin, by Vector Laboratories (1 mentions)
Digital optical microscope, by Leica camera (1 mentions)
Tcs sp laser scanning confocal microscope, by Leica camera (1 mentions)
Alexa 488, by Thermo Fisher Scientific (1 mentions)
Ab15093, by Abcam (1 mentions)
Ab13840, by Abcam (1 mentions)
Rl 1072, by Vector Laboratories (1 mentions)
Rhodamine labeled pna, by Vector Laboratories (1 mentions)
Dm400bled368424, by Leica camera (1 mentions)
Sc 28319, by Abcam (1 mentions)
24 protocols using image pro plus
1
Automated Immunofluorescence Imaging Protocol
2
Nerve Regeneration Histology Protocol
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The rabbits were euthanized at 12 weeks after operation via anesthetic overdose. The distal segment of regenerative
nerve was
harvested, followed by postfixing in 4% formaldehyde, embedding in
paraffin wax, and transverse sectioning at 3 μm. After blocking
with serum, sections were stained with H&E and primary antibodies
against neurofilament (NF, Gene Tex) and S100 (Gene Tex). The sections
were then incubated with goat anti-rabbit IgG antibodies (Invitrogen)
and exposed to hematoxylin to stain the cell nucleus. Finally, images
were acquired via a fluorescence microscope (Leica)
and analyzed using Image-Pro Plus.
nerve was
harvested, followed by postfixing in 4% formaldehyde, embedding in
paraffin wax, and transverse sectioning at 3 μm. After blocking
with serum, sections were stained with H&E and primary antibodies
against neurofilament (NF, Gene Tex) and S100 (Gene Tex). The sections
were then incubated with goat anti-rabbit IgG antibodies (Invitrogen)
and exposed to hematoxylin to stain the cell nucleus. Finally, images
were acquired via a fluorescence microscope (Leica)
and analyzed using Image-Pro Plus.
3
Quantifying Collagen and Vascular Profiles
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Slides were prepared as previously involved in H&E and Masson’s trichrome staining. Three sections from each group were performed for immunohistochemistry as described previously [17 (link)]. The primary antibodies, including antibodies against collagen 1A1 (Abcam, USA), collagen 3A1 (Abcam) and CD31 (Abcam) were used in immunohistochemistry. Digital images were obtained by microscope (Leica) and the intensity of each item was evaluated by Image-Pro Plus. Meanwhile, three slices of each group were stained with picrosirius red to assess the collagen volume fraction. Digital images were acquired with a polarized microscope. Collagen volume fraction for each group was evaluated by Image-Pro Plus software.
4
Immunofluorescence Staining of Cells
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Immunofluorescence was performed as previously described21 (link). Briefly, cells were seeded in 24-well dishes and fixed by 4% paraformaldehyde and stained with primary antibodies followed by FITC-conjugated secondary antibodies. Images were collected by fluorescent microscopy (Leica, Germany) and processed with ImagePro Plus.
5
Immunofluorescence Assay for EMT Markers
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The IF assay was performed according to previously established protocols (23 (link)). Cells were seeded into 24-well plates and fixed with 4% paraformaldehyde after 48-h treatment of puerarin or control. Fixed cells were stained with vimentin, Snail, and Slug, followed by incubation with FITC-conjugated anti-mouse IgG and FITC-conjugated anti-rabbit IgG (Cell Signaling Technology). Representative images were acquired by fluorescence microscopy (Leica), and the data were processed via ImagePro Plus (USA).
6
Immunofluorescence Imaging of Endometrial Cells
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Human endometrial stromal cells were fixed in 4% paraformaldehyde and then washed in phosphate-buffered saline (PBS). After permeabilization with 0.2% Triton X-100 in PBS (PBST), the cells were blocked in 1% bovine serum albumin/PBST. Primary antibody and secondary antibody incubations were performed for 60 min at room temperature in sequence. Nuclei were counterstained using 4′,6-diamidino-2-phenylindole (DAPI; Abcam). The staining was viewed using a microscope (DM6 B; Leica, Germany) and quantified by mean optical density using Image-Pro Plus. Five random fields per sample were used for counting. The antibodies used in this study are listed in Supplementary Table S2 .
7
Histological Quantification of Intimal Hyperplasia
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Extracted blood vessels were embedded and sectioned as described earlier, and stained with hematoxylin and eosin. All sections were observed under an optical microscope (DFC300FX; Leica Microsystems) for intimal hyperplasia and vascular wall damage. Data analysis was carried out by use of an Image-Pro Plus computer-assisted digital system (Leica Qwin Plus V3.2.1; Leica Microsystems). Stent area and the lumen area (LA) were calculated, and the neointimal area (NA) was calculated using the formula (strut area − luminal area). Percentage stenosis area was calculated as (1 − luminal area/strut area) ×100%.
8
Confocal and Electron Microscopy of Biofilm Granules
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CLSM images were recorded using a TCS SP confocal laser scanning microscope (Leica, Germany). Granules’ bacteria were stained using the SYTO 60 nucleic acid stain (Molecular Probes, Eugene, OR, USA). The stock solution of SYTO 60 as supplied was used at a dilution of 1:1000 in deionized water. Glycoconjugates of granules were stained with the Aleuria aurantia lectin (Vector, Burlingame, CA, USA) conjugated to Alexa488 (Molecular Probes) according to a screening of all commercially available lectins on the same type of biofilms [32 (link)]. Detailed information on the methods can be found elsewhere [33 ].
SEM were fixed with 2.5% glutaraldehyde in 0.1 M phosphate buffer (pH = 7.3) for up to 12 h at 4 °C and post-fixed in 1% OsO4 at room temperature for 90 min. Further SEM sample preparations and microscopic analysis were performed in our previous studies [12 (link),31 (link)].
Granule size, distribution of diameters, length and width of filamentous bacteria and roundness were analyzed using a Leica Digital optical microscope and Image-Pro Plus.
SEM were fixed with 2.5% glutaraldehyde in 0.1 M phosphate buffer (pH = 7.3) for up to 12 h at 4 °C and post-fixed in 1% OsO4 at room temperature for 90 min. Further SEM sample preparations and microscopic analysis were performed in our previous studies [12 (link),31 (link)].
Granule size, distribution of diameters, length and width of filamentous bacteria and roundness were analyzed using a Leica Digital optical microscope and Image-Pro Plus.
9
Immunofluorescence Analysis of p65 Localization
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Immunofluorescence analysis was processed according to protocols. Cells were implanted in 24-well dishes and fixed by 4% paraformaldehyde for 24h. Fixed cells were stained with p65 (1:200) (Cell Signaling Technology) followed by Cy3–conjugated anti–rabbit IgG (1:200) (abcam, USA). Representative images were detected by fluorescent microscopy (Leica, Germany) and data were processed via ImagePro Plus.
10
Histological analysis of mouse testes
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For histology study, mouse testes were fixed in Bouin's fixative at 4°C overnight for staining with hematoxylin and eosin as described previously (Wang et al., 2017 (link)). For IHF, testes were fixed with 4% paraformaldehyde in PBS at 4°C overnight and embedded in paraffin. The 4-μm-thick testicular sections were incubated with primary antibodies and washed three times with PBS containing 0.5% Tween 20. Nuclei were stained in 0.5 μg/mL DAPI after blotting with FITC- or TRITC-conjugated second antibodies. Slides were imaged under a fluorescent microscope (Leica, DM400BLED368424) or a Lecia TCS/SP5 confocal microscope and processed with Image-Pro Plus. Antibodies and fluorescent probes used in this study: PLZF (Santa Cruz Biotechnology, SC-28319), DDX4 (Abcam, ab13840), SYCP3 (Abcam, ab15093), PCNA (Abcam, ab29), Acrosin/ACR (Atlas Antibodies, HPA048687), rhodamine-labeled PNA (Vector Laboratories, RL-1072), Alexa Fluor 488- or TRITC-conjugated anti-mouse, or anti-rabbit secondary antibodies (Jackson ImmunoResearch).
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