Primary antibodies: mouse monoclonal anti-PML (Santa Cruz; PG-M3; 1:400), mouse monoclonal anti-PML (Millipore; clone 36.1-104; 1:400), rabbit monoclonal anti-K15 (Abcam; ab52816; 1:400), rabbit anti-K14 (Abcam; ab175549; 1:400), rabbit anti-K5 (Abcam; ab24647; 1:400), and K10 (Abcam; RKSE60; 1:200). Secondary antibodies used: Alexa Fluor® 488 goat anti-mouse IgG (H + L) (Molecular Probes; 1:500) and Alexa Fluor® 594 goat anti-rabbit IgG (H + L) (Molecular Probes; 1:500).
Ab52816
Manufactured by Abcam
Sourced in United Kingdom
Ab52816 is a laboratory product manufactured by Abcam. It serves as a general-purpose laboratory equipment. The core function of this product is to facilitate various experimental procedures and analyses conducted in research and scientific settings.
Automatically generated - may contain errors
Lab products found in correlation
Ab16667, by Abcam (3 mentions)
4 6 diamidino 2 phenylindole (dapi), by Solarbio (2 mentions)
Ab181551, by Abcam (2 mentions)
Accuri c6, by BD (2 mentions)
Ab32575, by Abcam (2 mentions)
Ab9026, by Abcam (2 mentions)
Ab6785, by Abcam (2 mentions)
Ab76318, by Abcam (2 mentions)
Alexa fluor 594 goat anti rabbit igg h l, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 488 goat anti mouse igg h l, by Thermo Fisher Scientific (1 mentions)
Ab24647, by Abcam (1 mentions)
Dm4000 microscope, by Leica (1 mentions)
Af761, by R&D Systems (1 mentions)
Mab386, by Merck Group (1 mentions)
Ab78078, by Abcam (1 mentions)
Ab18207, by Abcam (1 mentions)
Ab155279, by Abcam (1 mentions)
S8809, by Merck Group (1 mentions)
Ab175471, by Abcam (1 mentions)
Ab214488, by Abcam (1 mentions)
17 protocols using ab52816
1
Immunofluorescence Staining of Keratins
2
Hair Follicle Histology and Immunostaining
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For histology analysis, Hematoxylin and Eosin (H&E) staining were performed according to standard protocols with minor modification: sections were incubated for 5 min in hematoxylin and 30 s in eosin solutions. Image acquisition was performed on a Leica DM4000 microscope. Only follicles with a clear and complete structure were chosen for length analysis. The length of hair follicles was measured by ImageJ.
For immunostaining, the frozen sections were blocked in PBS with 5% Donkey serum or 1% BSA and 0.25% Triton for 1–4 h at room temperature, then incubated with primary antibody at 4 °C overnight and were subsequently incubated with secondary antibodies conjugated with Alexa Fluor 488, 594 or 647 (1:1000, Invitrogen, California, USA). Nuclei were stained with DAPI (Solarbio, Beijing, China). The following primary antibodies were used: P-cadherin antibody (1:1000, AF761, R&D), K15 (1:100, ab52816, Abcam), and Lef1 (1:200, 2230, CST). Image acquisition was performed on a Zeiss microscope. For quantification, 5 sections from each sample were processed and 5 randomly selected areas of each section were quantified in Image J.
For immunostaining, the frozen sections were blocked in PBS with 5% Donkey serum or 1% BSA and 0.25% Triton for 1–4 h at room temperature, then incubated with primary antibody at 4 °C overnight and were subsequently incubated with secondary antibodies conjugated with Alexa Fluor 488, 594 or 647 (1:1000, Invitrogen, California, USA). Nuclei were stained with DAPI (Solarbio, Beijing, China). The following primary antibodies were used: P-cadherin antibody (1:1000, AF761, R&D), K15 (1:100, ab52816, Abcam), and Lef1 (1:200, 2230, CST). Image acquisition was performed on a Zeiss microscope. For quantification, 5 sections from each sample were processed and 5 randomly selected areas of each section were quantified in Image J.
3
Immunolabeling for Myelination Markers
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The primary antibodies included Anti-beta III Tubulin [2G10] (1:1000, ab78078; Abcam, Cambridge, UK), Recombinant Anti-Cytokeratin 15 (1:200, ab52816; Abcam), Anti-beta III Tubulin (1:300, ab18207; Abcam), Recombinant Anti-SOX10 [EPR4007] (1:100, ab155279; Abcam), Monoclonal Anti-Myelin Basic Protein (MBP, 1:100, MAB386 Sigma-Aldrich), Monoclonal Anti-Sodium Channel, Pan antibody produced in mouse (1:500, S8809; Sigma-Aldrich), and polyclonal Anti-Caspr (1:1000, kindly provided by Elior Peles, PhD, Weizmann Institute of Science, Rehovot, Israel), Recombinant Monoclonal Anti-Mayelin Associated Glycoprotein [EPR24276] (MAG, 1:500, ab277524, Abcam).
4
Immunofluorescence Assay for Cytokeratin 15
5
Immunohistochemical Analysis of Prostate Cancer Markers
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Primary antibodies included ZEB1(21544‐1‐AP, Proteintech, 1:50), KLK3(ZM‐0218, ZSGB‐BIO), TACSTD2(ab214488, Abcam, 1:500), and KRT15(ab52816, Abcam, 1:200). Formalin‐fixed and paraffin‐embedded tissue sections (5 µm) were deparaffinized and rehydrated. Antigen retrieval was carried out using 10 mm sodium citrate (pH 6.0). Endogenous peroxidase activity was blocked with 0.3% hydrogen peroxide for10 min and 5% BSA in PBS for 1 h. Slides were incubated overnight at 4 °C with a primary antibody, which was followed by incubation with an HRP‐linked secondary antibody (PV‐9001, ORIGENE) at room temperature (30 min). Diaminobenzidine (DAB) was used as chromogen, and the sections were counterstained with haematoxylin. Representative fields were selected from these slides.
6
Immunohistochemical Analysis of Skin Markers
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The skin tissues were immersed in 4% paraformaldehyde (Sigma-Aldrich, St. Louis, MO, USA) for 30 min, subjected to gradient dehydration with ethanol, and embedded in paraffin. The resulting 6-μm skin sections were dewaxed with xylene, blocked with an IHC blocking buffer (Beyotime Biotechnology, Zhejiang, China) at 37°C for 30 min, and washed (three times, 5 min each, at room temperature) with an IHC washing buffer (Beyotime Biotechnology, China).
Next, the sections were incubated at 37°C for 45 min with primary antibodies against CD200 (ab244560), CD49F (ab181551), SOX-9 (ab185966), CK15 (ab52816), and FZD10 (ab137491) (Abcam, Cambridge, UK). Following another round of washing with an IHC washing solution (three times, 5 min each, at room temperature), the sections were incubated at 37°C for 45 min with the respective secondary antibodies. After the sections were washed thoroughly, they were blocked using an immunofluorescence (IF) blocking buffer (Sigma-Aldrich, USA).
Next, the sections were incubated at 37°C for 45 min with primary antibodies against CD200 (ab244560), CD49F (ab181551), SOX-9 (ab185966), CK15 (ab52816), and FZD10 (ab137491) (Abcam, Cambridge, UK). Following another round of washing with an IHC washing solution (three times, 5 min each, at room temperature), the sections were incubated at 37°C for 45 min with the respective secondary antibodies. After the sections were washed thoroughly, they were blocked using an immunofluorescence (IF) blocking buffer (Sigma-Aldrich, USA).
7
Immunophenotyping of Skin Cell Cultures
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All chemicals were obtained from Sigma-Aldrich (St. Louis, MO, USA) unless otherwise stated. Recombinant human IL-17A and IL-22 were purchased from PeproTech (Rocky Hill, NJ, USA). The following monoclonal antibodies (mAbs) were used: anti-BrdU antibody (BU-33; B2531) was obtained from Sigma-Aldrich, anti-BrdU antibody conjugated with FITC (ab74545) was purchased from Abcam (Cambridge, MA, USA), antibodies against differentiation-specific markers, namely K15 (ab52816; Abcam), K10 (SC31770; Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA), integrin β1/FITC (ab46920), filaggrin (ab24584), and Ki67 (ab16667) were all purchased from Abcam. Secondary antibodies conjugated with Alexa Fluor 488 (green; A11001) or 555 (red; A21432) were purchased from Molecular Probes (Eugene, OR, USA). Mowiol 4–88 anti-fade mounting solution was obtained from Polyscience, Inc. (Warrington, PA, USA).
8
Immunofluorescence Analysis of Primary Cell Markers
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Primary cells were washed with Phosphate-Buffered Saline (P1010, Solarbio, China) and fixed with 4% paraformaldehyde for 30 min, followed by treatment with 0.5% Triton X-100 for 20 min at room temperature. Next, the cells were blocked in 5% BSA (A8010, Solarbio, China) for 1 h at 37 °C and incubated with primary antibodies against Keratin (1:200, ab8068, Abcam, UK), K19 (1:200, PAB 30070, Bioswamp, China), and K15 (1:200, ab52816, Abcam, UK) at 4 °C overnight. The next day, the cells were incubated with Alexa Fluor 594-conjugated Goat Anti-Rabbit antibody (1:200, SAB43732, Bioswamp, China) and Alexa Fluor 594-conjugated Goat Anti-Mouse IgG (H+L) (1:200, SAB45897, Bioswamp, China) for 1 h at 37 °C and the nuclei were stained with 4’,6-diamidino-2-phenylindole (DAPI, S2110, Solarbio, China). Immunofluorescence was observed under a fluorescence microscope (DMIL LED, Leica, Germany).
9
Multimarker Immunofluorescence Analysis
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Anti‐Ki‐67 [SP6] (Abcam, ab16667) (1:50), Phospho‐Histone H3 (S10) (6G3) (Cell signalling, #9706) (1:100), Phospho‐Histone H3 (S10) (Abcam, ab5176) (1:100), Cleaved Caspase‐3 (Asp175) (Cell signalling #9661) (1:50), Anti‐Cytokeratin 15 [LHK15] (Abcam, ab80522) (1:500), Anti‐Cytokeratin 15 [EPR1614Y] (Abcam, ab52816) (1:500). Goat anti‐Mouse/Rabbit Secondary Antibodies, Alexa Fluor 488/594 (Invitrogen, #A11001, #A11005 #A11008 #A11037) (1:200). See also supporting references (Purba et al, 2016 , 2017a ,b ).
10
Characterizing ESCs via Immunostaining
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ESCs (1 × 106/ml) were trypsinized and suspended in 2% BSA/PBS (16000-044; Gibco) after 10 days of culture. After centrifuge and resuspension, the cells were incubated for 2 hours at room temperature with the following primary antibodies: anti-CK10 (1:100, ab9026; Abcam), anti-CK15 (1:100, ab52816; Abcam), and anti-α-SMA (1:20, ab32575; Abcam). Followed by centrifuge and washing, the resuspended cells were added in FITC-labeled secondary antibody IgG (1:500, ab6785; Abcam), and incubated for 30 minutes. The expression of CK10, CK15, and α-SMA was detected by BD Accuri C6 (BD, USA). Independent experiments were done in triplicate.
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