Western blot analyses were performed as described previously [37 (link)]. The protein concentrations of cell or tissue lysates were measured using the Lowry assay. Afterward, 40 μg protein samples were separated using 10% or 12% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE, according to the molecular weight of the proteins of interest) and then electroblotted onto a nitrocellulose membrane. The membranes were blocked with BlockPRO blocking buffer (BP01-1L; Visual Protein, Taipei, Taiwan), immunoblotted with specific primary antibodies against THBS2 (1:1000, A8561; ABclonal, Woburn, MA, USA), HSP90 (1:500, sc-101494; Santa Cruz Biotechnology), transferrin (1:1000, GTX112729; Gene Tex, Irvine, CA, USA), CD 81 (1:500, GTX101766; Gene Tex), α-SMA (1:1000, bsm-52396R; Bioss Antibodies Inc, Woburn, MA, USA), β-catenin (1:1000, GTX632676; Gene Tex), Fibronetin (1:500, ab2413; Abcam, Cambridge, MA, USA), Vimentin (1:1000, SC-7557; Santa Cruz Biotechnology), and β-actin (1:1000, NB600-501; Novus Biologicals, Centennial, CO, USA), and then detected using horseradish peroxidase-conjugated secondary antibodies. The signals were visualized by chemiluminescence using an enhanced detection kit (ECL, TU-ECL03, TOOLS, Taipei, Taiwan).
Sc 7557
Manufactured by Santa Cruz Biotechnology
Sourced in United States
Sc-7557 is a laboratory equipment product from Santa Cruz Biotechnology. It is designed for scientific research purposes.
Automatically generated - may contain errors
Lab products found in correlation
Ab2413, by Abcam (2 mentions)
A8561, by ABclonal (2 mentions)
Gtx632676, by GeneTex (2 mentions)
Sc 9068, by Merck Group (2 mentions)
Sc 25826, by Santa Cruz Biotechnology (2 mentions)
Sc 26643 r, by Santa Cruz Biotechnology (2 mentions)
Ax10 microscope, by Zeiss (2 mentions)
Sc 101494, by Santa Cruz Biotechnology (1 mentions)
Gtx101766, by GeneTex (1 mentions)
Nb600 501, by Novus Biologicals (1 mentions)
Eclipse e600 research microscope, by Nikon (1 mentions)
Tl 125 qhd, by Thermo Fisher Scientific (1 mentions)
Bsm 52396r, by Bioss Antibodies (1 mentions)
Sc 166362, by Santa Cruz Biotechnology (1 mentions)
Sc 2024, by Santa Cruz Biotechnology (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
Immpact dab kit, by Vector Laboratories (1 mentions)
Mab7119, by R&D Systems (1 mentions)
Mbs9406409, by MyBioSource (1 mentions)
Gtx112346, by GeneTex (1 mentions)
12 protocols using sc 7557
1
Western Blot Protein Analysis Protocol
2
Immunofluorescence Labeling of Paraffin-Embedded Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Sections (4 μm) were cut from paraffin embedded tissue and non-specific binding sites were saturated for 1 h at 37°C with PBS, 5% BSA. Sections were incubated at 4°C overnight with primary antibodies diluted in PBS supplemented with 1% BSA. The primary antibodies used were directed against VIM (Santa Cruz sc-7557), FN (Santa Cruz sc-9068), α-SMA (Sigma A5228), or HPSE (Santa Cruz sc-25826). Primary antibodies were revealed with an anti-goat-FITC for VIM, anti-rabbit-Cy3 for FN and HPSE, and anti-mouse-TR for α-SMA by incubation at room temperature for 45 min. Cell nuclei were visualized by Hoechst 33258. Images were obtained with a confocal LeicaSP5 microscope. Image processing was done with Image J (https://imagej.nih.gov/ij/ ).
3
Immunohistochemical Characterization of Cardiac Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The sections from cardiac tissues were examined using the ultravision quanto detection system procedure (TL-125-QHD, Thermo Fisher Scientific). The cardiac tissue sections were incubated with anti-THBS2 (1:200, A8561, ABclonal), α-SMA (1:400, bsm-52396R, Bioss Antibodies Inc), β-catenin (1:400, GTX632676, Gene Tex), fibronectin (1:200, ab2413, Abcam) and vimentin (1:400, SC-7557, Santa Cruz Biotechnology) antibodies at 4 °C overnight. Then, the sections were treated with a biotinylated anti-mouse secondary antibody for 1 h. The slides were counterstained with Mayer’s hematoxylin for 2 min and then mounted. The images were acquired using Nikon’s Eclipse E600 research microscope (Nikon, Tokyo, Japan).
4
ALDH and Vimentin Expression in DLD-S ALDH/PI CSCs
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The expression of ALDH and vimentin in DLD-S ALDH/PI CSCs were analyzed with immunofluorescence staining and the laser confocal technique. For immunofluorescence analysis, cells were grown in serum-free DMEM/F12 (Hyclone; GE Healthcare Life Sciences) medium containing several growth factors and placed onto glass-coverslips for 16 h prior to fixation with 2% paraformaldehyde at 37°C for 15 min. Fixed cells were then permeablized with 0.1% Triton X-100 in PBS at 20°C for 15 min, followed by incubation at 4°C overnight with the following primary antibodies: ALDH (1:200; sc-166362; Santa Cruz Biotechnology, Inc., Dallas, TX, USA) and vimentin (1:200; sc-7557; Santa Cruz Biotechnology), respectively. Antibodies were diluted using blocking buffer (PBS containing 2% BSA and 5% FBS). Cells were then incubated with phycoerythrin-(1:200; sc-166362; Santa Cruz Biotechnology, Inc.) or fluorescein isothiocyanate-conjugated secondary antibody (1:200; sc-2024; Santa Cruz Biotechnology, Inc.) at 20°C for 1 h. These slides were then stained using 4′,6-diamidino-2-phenylindole (DAPI; 1:1,000; Invitrogen; Thermo Fisher Scientific, Inc.) at 20°C for 10 min. Fluorescent images were captured using the Olympus FV1200 laser confocal microscope (Olympus Corporation, Tokyo, Japan).
5
HMGCR and HMGCS1 in TNBC Metastasis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Formalin-fixed, paraffin-embedded (FFPE) primary TNBC tumors and matched lung metastasis samples from four individuals with breast cancer were selected. The institutional review board (IRB) of Cedars-Sinai Medical Center and the First Hospital of China Medical University (Shenyang, China) granted approval. The normal human lung tissue samples were purchased from US Biomax (T041a). Primary mouse breast tumors and lung metastasis samples (see Mouse models and study approval ) were also used in the IHC assay. IHC staining was performed using the Vectastain ABC Kit (Vector Laboratories) and ImmPACT DAB Kit (Vector Labs). The primary antibodies were HMGCR (for human samples, 1:100, PA5-52547, Thermo Fisher Scientific), Hmgcr (for mouse samples, 1:70, MBS9406409, Mybiosource), HMGCS1 (for human and mouse samples, 1:100, GTX112346, GeneTex), CCR2 (1:100, NBP2-35334, Novus Biologicals), SREBP2 (1:100, MAB7119, R&D Systems), and Vimentin (1:100, sc-7557, Santa Cruz).
6
Immunohistochemical Localization of Vimentin, Fibronectin, α-SMA, and Heparanase
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Sections (4 μm) were cut from paraffin embedded tissue and non-specific binding sites were saturated for 1 h at 37°C with PBS, 5% BSA. Sections were incubated at 4°C overnight with primary antibodies diluted in PBS supplemented with 1% BSA. The primary antibodies used were directed against vimentin (Santa Cruz sc-7557), fibronectin (Santa Cruz sc-9068), α-SMA (Sigma A5228), or heparanase (Santa Cruz sc-25826). Primary antibodies were visualized with anti-mouse AlexaFluor-488, anti-goat-FITC, anti-rabbit-Cy3, and anti-mouse-TR secondary antibodies. Cell nuclei were visualized by Hoechst 33258. Images were obtained by a confocal LeicaSP5 microscope [35 (link)].
7
Immunohistochemical Profiling of BECs
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Sections were incubated with primary antibodies against TRPS1 (diluted 1:200; sc-26975), E-Cadherin (1:400; sc-1500), CK19 (1:200; sc-33119), Vimentin (1:200; sc-7557), α-SMA (1:200, sc-324317), and β-catenin (1:500; sc-81178; all from Santa Cruz Biotechnology, CA, USA). The secondary antibodies used were from an immunohistochemistry staining kit (Zhongshan Biotechnology Company, Beijing, China). After diaminobenzidine (DAB) staining, sections were counterstained with hematoxylin. For negative controls, primary antibodies were replaced with phosphate-buffered saline (PBS). The presence of brown granules was considered a positive signal. Five fields of view at high magnification were selected for each section and 100 BECs assessed in each field of view.
8
Immunohistochemical Analysis of Colon Tissues
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Colon tissues were fixed in 4% paraformaldehyde and embedded in paraffin. After incubation with blocking buffer, the sections were incubated with a goat polyclonal antivimentin antibody (sc-7557, 1:50 dilution; Santa Cruz Biotechnology, Santa Cruz, CA), a rabbit polyclonal anti-F4/80 antibody (sc-26643-R, 1:50 dilution; Santa Cruz Biotechnology), or a rabbit polyclonal anti-phospho-ERK antibody (no. 4370, 1:50 dilution; Cell Signaling Technologies, Danvers, MA) overnight at 4°C. After washing, the sections were incubated with donkey anti-goat IgG or bovine anti-rabbit IgG, and the slides were stained with an ABC kit for color development (sc-2018; Santa Cruz Biotechnology). Immunohistochemistry was assisted by Translational Pathology Core Laboratory (TPCL) of UCLA. Images were analyzed with a Zeiss AX10 microscope at magnification of 200×.
9
Immunofluorescence Staining of Cryosections
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cryosections were blocked in PBS containing 5% normal donkey serum for 2 h at room temperature. Sections were incubated over night at 4 °C with the primary antibodies against PDGFRβ (ab32570, rabbit Anti-PDGF Receptor beta antibody, 1:50, Abcam), Pan-Keratin (#4545, mouse anti-pan-keratin antibody,1:50, CST), Vimentin (sc-7557, goat anti-vimentin antibody, 1:50, Santa Cruz). After washed with PBS three times, the sections were incubated for 1 h at room temperature with Alexa Fluor 594-labeled donkey anti-rabbit IgG (A21207,1:400, Invitrogen), Alexa Fluor 488-labeled donkey anti-mouse IgG (A21202,1:400, Invitrogen) and Alexa Fluor 555-labeled rabbit anti-goat IgG (A21431,1:400, Invitrogen). Sections were washed three times in PBS, followed by mounting tissue with Dako fluorescence mounting medium. Photographs were taken using a Leica DMi8.
10
Immunostaining of Lung Tissue Sections
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!