Immunohistochemistry for cleaved caspase 3 was used to measure apoptotic rates in ex vivo tumor using a rabbit polyclonal anti-human antibody to cleaved caspase 3 (BioCare Medical, Concord, CA, #CP229B). Paraffin embedded tumor sections were heated, deparaffinized, and antigen retrieval was performed by steaming, and endogenous peroxides were blocked with 3% hydrogen peroxide in methanol. Non-specific proteins were blocked with 4% fish gelatin in PBS. Slides were incubated in primary antibody (1:100), and the secondary antibody (ready-to-use) was followed by streptavidin HRP (ready-to-use). Slides were quantified by counting the number of positively-staining cells per 200x field.
Cp229b
Manufactured by Biocare Medical
The CP229B is a centrifuge equipment designed for laboratory applications. It features a compact and durable construction, enabling efficient separation of samples in clinical, research, or diagnostic settings.
Automatically generated - may contain errors
Lab products found in correlation
Cleaved caspase 3, by Biocare Medical (1 mentions)
Ab156956, by Abcam (1 mentions)
Ab92547, by Abcam (1 mentions)
Ab16667, by Abcam (1 mentions)
Lotus tetragonolobus ltl, by Vector Laboratories (1 mentions)
Dolichos biflorus agglutinin, by Vector Laboratories (1 mentions)
Mca341r, by Bio-Rad (1 mentions)
Refine dab, by Leica (1 mentions)
5 protocols using cp229b
1
Measuring Apoptosis in Tumor Tissue
2
Histological Analysis of Drug Effects on Tumor-Bearing Mice
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For histologic analysis, brain tissues from control and drug-treated tumor-bearing mice were harvested, snap frozen in optimal cutting temperature embedding medium and stored at − 80°C. Cryostat sections were placed on slides and fixed in zinc-buffered formalin. Slides were blocked with 5% goat serum in 1% BSA followed by overnight incubation with primary antibodies against CD31 (102402, Biolegend, San Diego, CA), Ki-67 (ab156956, Abcam, Cambridge, MA), Vimentin (ab92547, Abcam), and cleaved-Caspase-3 (CP229B, Biocare Medical). Slides were then incubated with secondary antibodies conjugated to Alexa Fluor 488 or 635, washed and treated with 4′,6-diamidino-2-phenylindole as a nuclear counterstain. The detection fluorophores used were limited to those around the inherent fluorescence spectra of Doxo (λex = 480 nm, λem = 550 to 590 nm) [23] (link) to avoid bleed-through and enable co-detection of the drug with respect to certain antigens. Epifluorescence photomicrographs were captured at 100 × and 400 × magnification using an FV1000 confocal microscope (Olympus of America, Center Valley, PA) equipped with multi-Argon, 405, 559, and 635 diodes. Quantitative analysis was performed with Slidebook 5 software (Intelligent Imaging Innovations, Denver, CO).
3
Kidney and Heart Cell Proliferation and Apoptosis
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Cell proliferation was analyzed using an anti-Ki-67 monoclonal antibody (ab16667, ABCAM, Cambridge, UK). Negative control reactions had the primary antibody replaced with the diluent while positive control ones were performed in mouse spleen sections. Each kidney section had eight cortical and two medullary fields analyzed at 400 × magnification. Cell proliferation rate was calculated as the ratio cells with positive nuclei/total cells.
Apoptosis was evaluated with an anti-active caspase-3 antibody (CP229B, BioCare Medical, Concord, CA). Control reactions and selection of kidney fields followed the same protocol used for Ki-67. Each heart section had 10 random fields analyzed. Quantification was performed as the ratio positive cells/total cells.
Cyst origin was analyzed using the biotinylated lectins Dolichos biflorus agglutinin (DBA) and Lotus Tetragonolobus (LTL) (Vector Laboratories, Burlingame, CA; cat.# B-1035 and B-1325, respectively). Cysts were defined as having a diameter ≥ 50 μm59 (link). To better characterize the patterns, we classified the cysts as small (diameter ≥ 50 μm and ≤ 300 μm) or large (diameter > 300 μm). Quantification was carried out by counting the number of positive and negative large and small cysts at a 200 × magnification. The results were expressed in percentage, as the ratio number of positive cysts/total number of cysts.
Apoptosis was evaluated with an anti-active caspase-3 antibody (CP229B, BioCare Medical, Concord, CA). Control reactions and selection of kidney fields followed the same protocol used for Ki-67. Each heart section had 10 random fields analyzed. Quantification was performed as the ratio positive cells/total cells.
Cyst origin was analyzed using the biotinylated lectins Dolichos biflorus agglutinin (DBA) and Lotus Tetragonolobus (LTL) (Vector Laboratories, Burlingame, CA; cat.# B-1035 and B-1325, respectively). Cysts were defined as having a diameter ≥ 50 μm59 (link). To better characterize the patterns, we classified the cysts as small (diameter ≥ 50 μm and ≤ 300 μm) or large (diameter > 300 μm). Quantification was carried out by counting the number of positive and negative large and small cysts at a 200 × magnification. The results were expressed in percentage, as the ratio number of positive cysts/total number of cysts.
4
Histological Analysis of Lung Sections
5
Immunohistochemical Analysis of Lymph Nodes
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