Antibodies for SCYL1 (A6735) and Actin (A5441) were purchased from Abconal Technology (Wuhan, China) and Sigma-Aldrich (St. Louis, MO, USA), separately. Anti-Mouse (31,430) and anti-Rabbit (31,460) HRP-conjugated secondary antibodies were from ThermoFisher Scientific (Waltham, MA, USA). The shRNA oligos for SCYL1 were produced by Sangon Biotech Company (Shanghai, China). Transwell chambers (3422) were purchased from Corning Inc (Corning, NY, USA). Puromycin was purchased from Biotopped (Beijing, China). The DAB kit for IHC was from Zhongshan Jinqiao Biotechnology (Beijing, China). The SABC-POD kit and Mayor’s hematoxylin were purchased from Boster Biological Technology (Wuhan, China). Human breast cancer cell lines MDA-MB-231 and MCF-7 were obtained from ATCC (Manassas, VA, USA).
Sabc pod kit
Manufactured by Boster Bio
Sourced in China
The SABC-POD kit is a laboratory equipment product designed for protein detection and quantification. It provides a reliable method for the analysis of target proteins in biological samples. The kit includes all the necessary components to perform the assay, enabling researchers to efficiently measure protein levels in their experiments.
Automatically generated - may contain errors
Lab products found in correlation
Transwell chambers 3422, by Corning (1 mentions)
Mcf 7, by American Type Culture Collection (1 mentions)
Mda mb 231, by American Type Culture Collection (1 mentions)
Ab183218, by Abcam (1 mentions)
Ab201792, by Abcam (1 mentions)
Ab133357, by Abcam (1 mentions)
Masson s trichrome staining, by Merck Group (1 mentions)
Sc 20047, by Santa Cruz Biotechnology (1 mentions)
Sc 377009, by Santa Cruz Biotechnology (1 mentions)
Ab9722, by Abcam (1 mentions)
Hematoxylin, by Solarbio (1 mentions)
Ab211542, by Abcam (1 mentions)
Peroxidase substrate kit, by Boster Bio (1 mentions)
Nb500 170, by Santa Cruz Biotechnology (1 mentions)
3 3 diaminobenzidine dab kit, by Boster Bio (1 mentions)
Nbp2 13740, by Proteintech (1 mentions)
He staining kit, by Solarbio (1 mentions)
Ab27988, by Abcam (1 mentions)
Diaminobenzidine dab, by Boster Bio (1 mentions)
9 protocols using sabc pod kit
1
Knockdown of SCYL1 in Breast Cancer
2
Comprehensive Immunohistochemical Profiling
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Immunohistochemistry (IHC) staining was performed according to a previous method [13 (link), 20 ]. Serial paraffin sections were subjected to antigen retrieval, incubation in antigen retrieval solution for 20 min, inactivation with endogenous peroxidase (3% H2O2), and blocked in goat serum for 1 h. Sections were then incubated with primary antibodies against CD3 (SC-20047, Santa Cruz Biotechnology Inc., Dallas, TX, USA), F4/80 (SC-377009, Santa Cruz Biotechnology Inc.), IL-6 (GeneTex, Santa Cruz Biotechnology Inc.), IL-1β (ab9722, Abcam), TNF-α (ab183218, Abcam), p-p65 (Ser536) (#3033, Cell Signaling Technology), p16 (ab211542, Abcam), NLRC4 (ab201792, Abcam), ITGAM (ab133357, Abcam), p19 (ab80, Abcam), Collagen I (14695-1-AP, Proteintech), Collagen III (22734-1-AP, Proteintech), α-SMA (14395-1-AP, Proteintech), and β-galactosidase (15518-1-AP, Proteintech). After washing, sections were incubated with a secondary antibody for 1 h, and processed using the SABC-POD kit (SA2001, Boster, China). Then, sections were counterstained with Hematoxylin and fixed with biomount medium. Hematoxylin and Shandon Instant Eosin (Solarbio Co., Ltd.) were used to determine cell infiltration. Masson's trichrome staining (Sigma-Aldrich®) was used to assess collagen deposition.
3
Quantification of IgA-Positive Cells in Tissue
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Immunohistochemical staining was performed with an SABC-POD Kit (Boster Biological Technology Co. Ltd.) according to the instructions of the manufacturer. Briefly, paraffin-embedded tissue sections were deparaffinized, rehydrated, and quenched with 3% hydrogen peroxide solution for 15 min at room temperature. The slides were pretreated by heating the slides for 25 min in 10 mM citrate buffer. After they had been washed with PBS, the slides were blocked with goat serum for 30 min. Rabbit Anti-Mouse IgA (GENXPAN) was used as the primary antibody, horseradish peroxidase (HRP) Goat Anti-Rabbit IgG (TransGen Biotech) was used as the secondary antibody. Tissue staining was visualized with 3,3′-diaminobenzidine (DAB). Images were captured using a microscope (Olympus, Tokyo, Japan). Eight disconnected slices were randomly selected at each time point, and 5 fields of view were selected for each slice for counting, and cells were normalized to field area (mm2). Mechanical Manual Cell Counter (YAMI, China) was used for cell counting. All evaluations were performed by two pathologists who were blinded to the treatment regimens.
4
Immunohistochemical Analysis of Enteric Viruses
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Briefly, paraffin sections were dewaxed and rehydrated as described above. Antigen retrieval was performed by incubating sections in 10 mM citrate buffer (pH 6.0) in a decloaking chamber for 30 min at 95°C. Sections were blocked with 5% normal goat serum and then incubated with anti-PDCoV-N, MUC-2, Notch-1, or HES-1 (1:100) overnight at 4°C in a humidified chamber. For negative controls, sections were incubated in buffered saline with Tween 20 (PBST) only. An SABC-POD kit (rabbit or mouse IgG) and a peroxidase substrate kit (both from BOSTER, Wuhan, China) were used, respectively, for amplification and visualization of signal.
5
Immunohistochemical Profiling of Tumor Markers
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All specimens collected were routinely embedded in paraffin, and the tissue slides were deparaffinized and rehydrated. H&E staining was performed with an H&E staining kit (Solarbio, G1120-100). For IHC staining, after incubation with 3% H2O2, antigen retrieval, and blocking, sections were incubated with primary anti-PCIF1 (Novus, NBP2-13740), anti-CTBP2 (Proteintech, 10346), anti-TET2 (Abcam, ab94580), anti-Ki67 (Novus, NB500-170), and anti-PCK (pan-cytokeratin; Santa Cruz, sc-8018) overnight at 4°C. After washing, secondary antibodies and streptavidin-biotin complex (SABC) were applied with an SABC-POD kit (BOSTER, SA1022). Then the slides were stained with a 3,3′-diaminobenzidine (DAB) kit (BOSTER) and counterstained with hematoxylin. For the IHC analysis, the IHC score was calculated by multiplying the proportion of positively stained cells at each intensity level by their respective intensity score (intensity scores ranging from 0 to 3). The final score ranges from 0 to 300. For Ki67, the percentage of positively stained cells was calculated. The anti-PCK antibody (Santa Cruz Biotechnology, sc-8018) was used for IHC staining of metastatic cells in cervical lymph nodes.
6
Immunohistochemical Evaluation of Cell Markers
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IHC was performed with primary antibodies against CD105 (A19008, ABclonal, China), CD44 (A12410, ABclonal, China), CPT1A (#12252, CST, USA), pan-cytokeratin (AE1/AE3) (ab27988, Abcam, UK), IL-8 (A2541, ABclonal, China) and STC1 (20621-1-AP, Proteintech, China) using instant SABC-POD Kit (Boster Biological Technology) following the manufacturer’s procedure. The samples were scanned and imaged using the Automatic digital slice scanning system. The staining scores of target proteins were obtained by the percentage of positive cells (< 5%: 0, 5–25%: 1, 26–50%: 2, 51–75%: 3, 76–100%: 4) multiplied by the staining intensity of positive cells (weak: 1, moderate: 2, strong: 3).
7
In Situ Hybridization of miR-361-3p in PDAC
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Human PDAC tissues were fixed in 4% paraformaldehyde (pH 7.0–7.6, containing 0.1% DEPC) and then embedded in paraffin. A digoxigenin-labeled miR-361-3p oligonucleotide probe (5′-AAATCAGAATCACACCTGGGGGA-3′) and an ISH kit (MK10503) were obtained from Boster Biological Technology (Wuhan, China). Slides were deparaffinized and incubated for 20 min at room temperature with Pepsin; subsequently, the sections were prehybridized in a humid chamber at 38–42 °C for 2 h. Then, the tissues were hybridized overnight with a miR-361-3p probe at 38-42 °C. After hybridization, the slides were washed with graded-diluted sodium citrate buffer (SSC) at 37 °C for 40 min, followed by incubation with an antibody against digoxigenin at 37 °C for 60 min. These sections were then incubated with SABC-POD kit (Boster, Wuhan, China), and hybridization signals were visualized using diaminobenzidine (DAB) (Boster, Wuhan, China). Finally, the tissues were counterstained with hematoxylin, mounted and imaged.
8
Immunohistochemical Staining Protocol
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Immunohistochemical staining was performed according to the manufacturer’s instructions for SABC-POD Kit (Boster, SA2002, Wuhan, China). Briefly, paraffin-embedded sections were first deparaffinized and rehydrated, and 3% H2O2 was used to block the endogenous peroxidase activity for 20 min at RT. The slides were washed with PBS, before the antigen retrieval was performed with Citrate Buffer (pH = 6.0) under high pressure. Blocking buffer (10% donkey serum in PBS + 0.1% Triton X-100, if a permeabilization was needed) was used to block the tissues sections for 1 h at RT. Primary antibodies, diluted in PBS with 1% donkey serum, were incubated on the slides at 4 °C overnight. After three washes with PBS, slides were then incubated with the biotin-labeled secondary antibody and SABC in turn at RT for 30 min. Finally, the slides were developed with color by DAB and counterstained with hematoxylin (if needed).
9
METTL3 Immunohistochemistry Staining
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IHC staining was performed using a SABC-POD Kit (Boster, Wuhan, China) according to the manufacturer's instructions. Brie y, slides were incubated with primary antibodies against METTL3 (1:250, Abcam, Cambridge, UK) at 37°C overnight and then treated with anti-rabbit secondary antibody at room temperature for 2h. Slides were then stained with DAB and counterstained with Mayer's hematoxylin. The number of positively stained cells and the intensity of positive staining on cells were independently scored by two pathologists in a blinded manner. As previously described [25] (link), the protein expression level was evaluated by a semi-quantitative immunoreactivity scoring system.
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