Cells were fixed with ice-cold methanol and incubated with 0.5% Triton. Unspecific epitops were blocked using 20% AB serum. Next, cells were incubated with their respective primary antibody or isotype control (mouse monoclonal anti-CXCR4, 1 : 250 dilution; Abcam; rabbit polyclonal anti-CXCR7, 1 : 500 dilution; GeneTex; mouse IgG1k (MOPC-21); 2 μg ml−1; Abcam; rabbit immunoglobulin fraction (Code X0903) 2 μg ml−1; Dako). After 45 min of incubation, cells were washed and treated with either anti-mouse- or anti-rabbit-Alexa Fluor 488 (10 μg ml−1; Thermo Fisher). Nuclear staining was achieved using DAPI (200 ng ml−1). After fixation with 1% PFA, the cells were visualised using a fluorescence microscope at × 400 magnification (Zeiss Axioplan 2, Carl Zeiss).
Rabbit polyclonal anti cxcr7
Manufactured by GeneTex
Sourced in United Kingdom
Rabbit polyclonal anti-CXCR7 is an antibody product developed by GeneTex. The antibody is raised in rabbits and recognizes the CXCR7 protein. CXCR7 is a chemokine receptor that binds to the chemokine CXCL12. This antibody can be used for the detection and analysis of CXCR7 in various applications.
Automatically generated - may contain errors
Lab products found in correlation
Mouse monoclonal anti cxcr4, by Abcam (5 mentions)
Mouse igg1k mopc 21, by Abcam (3 mentions)
Alexa fluor 488 anti rabbit, by Thermo Fisher Scientific (2 mentions)
Rabbit immunoglobulin fraction code x0903, by Agilent Technologies (2 mentions)
Axioplan 2, by Zeiss (2 mentions)
Biophotometer, by Eppendorf (2 mentions)
5 protocols using rabbit polyclonal anti cxcr7
1
Immunofluorescence Staining of Chemokine Receptors
2
Protein Isolation and Immunoblotting
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For protein isolation, cells were lysed in ice-cold RIPA buffer. Total protein concentrations were measured using a BioPhotometer (Eppendorf, Hamburg, Germany). Proteins were separated using SDS–polyacrylamid gel electrophoresis and transferred onto a nitrocellulose membrane for antibody-detection. Membranes were incubated overnight using the following primary antibodies: mouse monoclonal anti-CXCR4 (1 : 100 dilution; Abcam), rabbit polyclonal anti-CXCR7 (1 : 200 dilution; GeneTex).
3
Immunofluorescence Analysis of CXCR4 and CXCR7
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Cells were cultivated in 6-well chamber slides and fixated with ice-cold methanol. After treatment with 0.5 % Triton the cells were blocked with 20 % AB serum and incubated with their respective primary antibody and isotype control (mouse monoclonal anti-CXCR4, 1:250 dilution; Abcam, Cambridge, UK; rabbit polyclonal anti-CXCR7, 1:500 dilution; GeneTex, Irvine, CA, USA; mouse IgG1k (MOPC-21); 2 µg/ml; Abcam, Cambridge, UK; rabbit immunoglobulin fraction (Code X0903) 2 µg/ml; Dako, Glostrup, Denmark). After 45 minutes of incubation, cells were washed and incubated with anti-mouse- or anti-rabbit-Alexa Fluor® 488 (10 µg/ml; Thermo Fisher, MA, USA), respectively. Nuclear staining was carried out with DAPI (200 ng/ml). After fixating the cells with 1 % PFA, visualization was achieved using a fluorescence microscope at 400x magnification (Zeiss Axioplan 2, Carl Zeiss, Göttingen, Germany).
4
CXCR4 and CXCR7 Protein Detection
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Cells were harvested using ice-cold RIPA buffer. Total protein concentrations were determined using a BioPhotometer® according to Bradford (Eppendorf, Hamburg, Germany). Protein separation was achieved by SDS-polyacrylamid gel electrophoresis and subsequent transfer onto a nitrocellulose membrane. Antibody-detection was performed overnight using the following primary antibodies: mouse monoclonal anti-CXCR4 (1:100 dilution; Abcam, Cambridge, UK), rabbit polyclonal anti-CXCR7 (1:200 dilution; GeneTex, Irvine, CA, USA).
5
Immunohistochemical Analysis of CXCR4 and CXCR7 in Follicular Thyroid Carcinoma
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Formalin-fixed paraffin-embedded tissue samples were obtained from the Institute of Pathology, University Hospital Duesseldorf. All tissue samples were newly reviewed prior to this study and the diagnosis of FTC histologically confirmed. FTCs were defined as thyroid carcinomas with follicular differentiation in the absence of papillary nuclear features, with either capsular or vascular invasion. The construction of the tissue microarray, immunohistochemistry and protein expression analyses were performed as described previously21 (link),22 (link). The expression levels of CXCR4 and CXCR7 were scored by two independent investigators (TW, CF) using the immunoreactivity score (IRS) reported by Remmele23 (link) in a blinded manner.
The immunohistochemistry was carried out using mouse monoclonal anti-CXCR4 (1:100 dilution; Abcam, Cambridge, UK) and rabbit polyclonal anti-CXCR7 (1:200 dilution; GeneTex, Irvine, CA, USA) as primary antibodies. Isotype control was performed using mouse IgG1k (MOPC-21; 1:50 dilution; Abcam, Cambridge, UK) and rabbit immunoglobulin fraction (Code X0903; 1:1000 dilution; Dako, Glostrup, Denmark). CXCR4 expressing tonsil tissue and CXCR7 expressing pancreatic adenocarcinoma served as positive controls. The prognostic power of CXCR4 and CXCR7 was assessed in accordance with the REporting recommendations for tumour MARKer prognostic studies (REMARK)24 .
The immunohistochemistry was carried out using mouse monoclonal anti-CXCR4 (1:100 dilution; Abcam, Cambridge, UK) and rabbit polyclonal anti-CXCR7 (1:200 dilution; GeneTex, Irvine, CA, USA) as primary antibodies. Isotype control was performed using mouse IgG1k (MOPC-21; 1:50 dilution; Abcam, Cambridge, UK) and rabbit immunoglobulin fraction (Code X0903; 1:1000 dilution; Dako, Glostrup, Denmark). CXCR4 expressing tonsil tissue and CXCR7 expressing pancreatic adenocarcinoma served as positive controls. The prognostic power of CXCR4 and CXCR7 was assessed in accordance with the REporting recommendations for tumour MARKer prognostic studies (REMARK)24 .
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