Immunohistochemistry was performed using the Double-Staining Methods based on Streptavidin Alkaline Phosphatase and Streptavidin Peroxidase labeling techniques (KIT-9999, MXB-China). All of the procedures were carried out according to the manufacturer's instructions. Sections were stained with anti-rabbit CD44 antibody (1:100, ZA-0537, ZSGB, China) and anti-mouse CD24 antibody (1:100, ab31622, Abcam, USA). To control the reliability of the CD44 and CD24 double-staining, single staining with CD44 and CD24 was done. Immunohistochemistry for AR (1:200, ZA-0554, ZSGB, China) and Ki67 (working solution, ZA-0502, ZSGB, China) were carried out as previously reported [36 (link)]. Sections were incubated with goat serum for negative controls of immunoreactions.
Za 0502
Manufactured by ZSGB-BIO
Sourced in China
The ZA-0502 is a laboratory centrifuge device. It is designed to separate components of a liquid mixture based on their differences in density, size, and shape. The core function of the ZA-0502 is to provide a controlled centrifugal force to facilitate the separation process.
Automatically generated - may contain errors
Lab products found in correlation
Anti ki67, by ZSGB-BIO (2 mentions)
Female nude mice, by Charles River Laboratories (2 mentions)
Ab137785, by Abcam (2 mentions)
Ab31622, by Abcam (1 mentions)
Kit 9999, by Beijing Strong Biotechnologies (1 mentions)
Ab28512, by Abcam (1 mentions)
Hematoxylin, by Absin (1 mentions)
Pv 9000 kit, by ZSGB-BIO (1 mentions)
Zli 9067, by ZSGB-BIO (1 mentions)
Zm 0069, by ZSGB-BIO (1 mentions)
Za 0550, by ZSGB-BIO (1 mentions)
Zli 9610, by ZSGB-BIO (1 mentions)
Sp 9001, by ZSGB-BIO (1 mentions)
9 protocols using za 0502
1
Multi-Marker Immunohistochemical Staining
2
Thyroid Tissue Immunohistochemistry Analysis
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Dissected thyroid and tumour tissues were formalin-fixed paraffin-embedded. Anti-Ki67 (ready-to-use, ZA-0502; Zsbio) was used for immunohistochemistry (IHC). For assessment of the intensity, each field was graded semiquantitatively on a three-tier scale (0 = negative staining, 1 = weak staining, 2 = moderate staining, 3 = strong staining). Histology and IHC scores were evaluated blindly by two independent pathologists (Dr. X.C. and Dr. Z.L.).
3
Immunohistochemistry Scoring of HCC Markers
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Immunohistochemistry analysis of AP4 (ab28512; Abcam, Cambridge, MA, USA), LAPTM4B (bs‐6542R Bioss, Boston, MA, USA) and Ki‐67 (ZA‐0502; ZSGB‐BIO, Beijing, China) was performed in HCC tissues according to a previous description (Li et al., 2017 ). Stained tissue sections were evaluated separately by two pathologists without any knowledge of the clinical data. For cytoplasmic staining, scoring was assessed based on the sum of cytoplasm staining intensity and the percentage of positive staining areas of cells. The staining intensity was scored as 0 (negative), 1 (weak), 2 (medium) and 3 (strong) and the percentage of positive staining areas of cells was defined as a scale of 0–3, where 0 represents < 10%, 1 was 10–25%, 2 was 26–75% and 3 was ≥ 76%. For nuclear staining, the score was defined according to the sum of nuclear staining intensity and numbers of cells. Nuclear staining intensity score was consistent with cytoplasm and positive nuclear staining scores were defined as follows: 0 represents < 10%, 1 was 10–50%, 2 was 51–80% and 3 was ≥ 80%. For statistical analysis, scores of 0–3 or 4–6 was, respectively, considered to be low or high expression.
4
Immunohistochemical Staining of FFPE Tumor Tissues
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Formalin-fixed, paraffin-embedded (FFPE) blocks of tumor tissue were sliced into 5 µm sections, deparaffinized in xylene (3×5 minutes) and then rehydrated through graded concentrations of ethanol (2×2 minutes in 100% ethanol, 1×2 minute in 95% ethanol, 1×2 minute in 75% ethanol). Tissue sections were then heated in EDTA solution (ZLI-9067; ZSGB-BIO, Beijing, China), for 150 seconds. After cooling, the sections were washed with PBS and then incubated with primary antibodies at 4°C overnight. The primary antibodies used were rabbit antibodies against CD34 (ZA-0550; ZSGB-BIO), CD99 (ZA-0577; ZSGB-BIO), Bcl-2 (ZA-0536; ZSGB-BIO), Ki67 (ZA-0502; ZSGB-BIO), and S100 (ZA-0225; ZSGB-BIO), as well as mouse antibodies against CK5/6 (ZM-0313; ZSGB-BIO) and CK-pan (ZM-0069; ZSGB-BIO). Slides were washed again and incubated with secondary antibodies using a PV-9000 kit (ZSGB-BIO) according to the manufacturer’s manual. The slides were then stained with hematoxylin (Absin, Fuzhou, China).
5
Immunohistochemical Analysis of ANXA3 and Ki67
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6
Immunohistochemical Staining for Cancer Markers
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After tumor tissue was taken, fixed and embedded with the paraffin and sectioned, the sections were dewaxed in xylene and rehydrated using graded concentrations of ethanol and distilled water. For HE assays, the sections were directly stained with hematoxylin-eosin. For IHC experiments, the procedures are described in the previous published article [10 (link)]. Briefly, the primary antibody was incubated overnight at 4 °C and the second antibody was incubated at room temperature for 30 min. The primary antibodies used in this article includes anti-YB1(#4202, CST, 1:50 dilution), anti-BRD7 antibody (51009–2-AP, proteintech, 1:500 dilution), anti-Ki67 antibody (ZA0502, ZSGB-BIO), anti-E-cadherin antibody (#24E10, CST, 1:100dilution) and anti-Vimentin antibody (ARG66302, arigo,1:500).
7
Subcutaneous Breast Cancer Xenograft Model in Nude Mice
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In vivo experiments were performed using female nude mice (4- to 5-week-old) purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. The animals were kept in a temperature- and humidity-controlled facility with a 12-h light/dark cycle with free access to food. Animal experiments were approved by the Sun Yat-sen University Cancer Center ethics committee. To establish a breast cancer model, 1 × 105 MDA-MB-231 cells (0.1 ml) were injected subcutaneously (five mice in each group). Every 3-4 days, tumor size was measured, and body weight was recorded. On the 39th day postinjection, the mice were sacrificed, and the tumors and organs were carefully removed, weighed, and subjected to immunohistochemical staining for Ki67 (ZA-0502, ZSGB-Bio, China) and PFKFB4 (ab137785, Abcam, USA). Tumor volumes were calculated with the following formula: A × B2/2 (A: the longest diameter; B: the diameter perpendicular to A).
8
Immunohistochemical Analysis of Ki-67 in Brain Tumors
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Brain tumors were immersed in 4% formalin, embedded in paraffin, cut into slices, and dewaxed. After antigen retrieval, the slides were blocked with BSA (bovine serum albumin) for 30 min at 37°C and then incubated with primary antibodies against Ki-67 (Zsbio, ZA-0502, 1:100 dilution) at 4°C overnight. Afterwards, the slides were rewarmed at room temperature for 1 h and then incubated with a biotin-labeled secondary antibodies (Zsbio, SP-9001). The DAB (diaminobenzidine) kit (Solarbio, DA1015) was used for staining, and hematoxylin was used for counterstaining. Finally, the samples were dehydrated, and the images were acquired by microscopy (Olympus, Japan).
9
Breast Cancer Xenograft Model in Nude Mice
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In vivo experiments were performed using female nude mice (4-to 5-week-old) purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. The animals were kept in a temperature-and humiditycontrolled facility with a 12-h light/dark cycle with free access to food. Animal experiments were approved by the Sun Yat-sen University Cancer Center ethics committee. To establish a breast cancer model, 1 × 10 5 MDA-MB-231 cells (0.1 ml) were injected subcutaneously ( ve mice in each group). Every 3-4 days, tumor size was measured, and body weight was recorded. On the 39th day post injection, the mice were sacri ced, and the tumors and organs were carefully removed, weighed and subjected to immunohistochemical staining for Ki67 (ZA-0502, ZSGB-Bio, China) and PFKFB4 (ab137785, Abcam, USA). Tumor volumes were calculated with the following formula: A × B 2 /2 (A: the longest diameter, B: the diameter perpendicular to A).
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