Podocalyxin-Like (PODXL) antibody

Details for Product No. ABIN967659
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Antigen
Synonyms PODXL, podxl, fc18d02, wu:fc18d02, Gp200, PC, PCLP, PCLP-1, AW121214, Ly102, Pclp1, podocalyxin, Gp135, PCLP1, cPCLP1, MEP21, Podxl
Reactivity
Human
(97), (25), (18), (1)
Host
Mouse
(52), (47), (8), (5), (2)
Clonality (Clone)
Monoclonal ()
Conjugate
Un-conjugated
(10), (8), (6), (4), (2), (2), (2), (1), (1), (1), (1), (1), (1), (1), (1), (1), (1)
Application
Western Blotting (WB), BioImaging (BI), Flow Cytometry (FACS), Immunocytochemistry (ICC), Immunoprecipitation (IP), Radioimmunoassay (RIA)
(83), (41), (29), (20), (12), (12), (12), (10), (10), (5), (2), (1), (1)
Pubmed 7 references available
Quantity 0.1 mg
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Catalog No. ABIN967659
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Immunogen Human Embryonal Carcinoma Cell Line
Clone TRA-1-60
Isotype IgM
Cross-Reactivity Rhesus Monkey
Characteristics 1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
2. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
3. Please refer to us for technical protocols.
4. This antibody has been developed and certified for the bioimaging application. However, a routine bioimaging test is not performed on every lot. Researchers are encouraged to titrate the reagent for optimal performance.
Purification Purified from tissue culture supernatant or ascites by affinity chromatography.
Alternative Name TRA-1-60
Background The TRA-1-60 monoclonal antibody reacts with the neuraminidase-resistant form of a pluripotent-stem-cell-specific epitope on a high-molecular-weight transmembrane glycoprotein. The TRA-1-60 antigen is a sialylated epitope on the same keratan sulfate core molecule, podocalyxin, as 4 other distinct antigens on tumor-derived cell lines, TRA-1-81, GCTM2, K4, and K21. The expression of TRA-1-60 antigen is stage-specific and can be used to characterize embryonic cells and monitor their differentiation. The antigen is found on teratocarcinoma (embryonal carcinoma or EC), embryonic inner cell mass (but not morula or trophoblast), and embryonic stem (ES) cells. TRA-1-60 antigen is released into the serum of patients bearing testicular tumors containing EC cells. As human EC and ES cells undergo differentiation, expression of TRA-1-60 antigen is lost. Expression of TRA-1-60 antigen has also been observed on a rhesus monkey ES cell line (Thomson et al, 1995).
Synonyms: TRA-1-60(R)
Molecular Weight Multiple, >200 kDa
Application Notes Bioimaging:
1. Seed the cells in appropriate culture medium at an appropriate cell density in an 96-well Imaging Plate , and culture overnight to 48 hours.
2. Remove the culture medium from the wells, wash the wells twice with 100 myl of 1× PBS, and fix the cells by adding 100 µl of fresh 3.7% Formaldehyde in PBS or fixation buffer to each well and incubating for 10 minutes at room temperature (RT).
3. Remove the fixative from the wells, and wash the wells twice with 100 myl of 1× PBS.
4. Dilute the antibody in 1× PBS, and stain the cells by adding 50 µl of the diluted antibody to each well and incubating for 1 hour at RT.
5. Remove the diluted antibody, and wash the wells three times with 100 myl of 1× PBS.
6. Remove the PBS, dilute the second-step reagent in 1× PBS, and stain the cells by adding 50 µl of the diluted second-step reagent to each well and incubating for 1 hour at RT.
7. Remove the diluted second-step reagent, and wash the wells twice with 100 myl of 1× PBS.
8. Remove the PBS, and counter-stain the nuclei by adding 100 ml of a 2 mg/ml solution of Hoechst 33342 in 1× PBS to each well at least 15 minutes before imaging.
9. View and analyze the cells on an appropriate imaging instrument.
Comment

Related Products: ABIN967389

Restrictions For Research Use only
Format Liquid
Concentration 0.5 mg/ml
Buffer Aqueous buffered solution.
Preservative Sodium azide
Precaution of Use This product contains sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
Storage 4 °C
Supplier Images
anti-Podocalyxin-Like (PODXL) antibody Immunofluorescent staining of human ES cell line. The H9 cell line (WiCell, Madison, WI) was cultured, fixed, and stained with purified mouse anti-human TRA-1-60 Antigen monoclonal antibody (clone TRA-1-60) (pseudo-colored green) according to the Recommended Assay Procedure. The second step reagent was Alexa Fluor® 647 goat anti-mouse Ig (Invitrogen). The left image shows the plasma membrane staining by the TRA-1-60 mAb, and the right image shows TRA-1-60 with counter-staining of the nuclei by Hoechst 33342 (pseudo-colored blue). The images were captured on a BD Pathway™ 435 Cell Analyzer using a 10X objective and merged using BD Attovision™ software. If permeabilization is required for staining additional markers, we recommend the use of cold 90% methanol or BD™ Phosflow Perm Buffer III.
anti-Podocalyxin-Like (PODXL) antibody (2) Immunofluorescent staining of human ES cell line
anti-Podocalyxin-Like (PODXL) antibody (3) Western Blot analysis of TRA-1-60 in mouse and human ES cell lines. Lysates from ES-E14TG2a mouse ES cells (ATCC CRL-1821, left blot) and H9 human ES cells (WiCell, Madison, WI, right blot) were probed with purified mouse anti-human TRA-1-60 Antigen monoclonal antibody at titrations of 2.0 (lanes 1), 1.0 (lanes 2), and 0.5 µg/ml (lanes 3). High-molecular-weight molecules bearing the TRA-1-60 epitope are identified above 200 kDa in the human ES cells. Expression of the TRA-1-60 epitope has not been reported in mouse ES cells.
The H9 cells were cultured on a mitomycin C-treated mouse embryonic fibroblast feeder layer [MEF (CF-1), ATCC SCRC-1040] that maintains the undifferentiated state of the ES cells. The lysate was made from a mixture of the 2 cell types, the majority of which were H9 cells.
Product cited in: Andrews, Banting, Damjanov et al.: "Three monoclonal antibodies defining distinct differentiation antigens associated with different high molecular weight polypeptides on the surface of human embryonal carcinoma cells." in: Hybridoma, Vol. 3, Issue 4, pp. 347-61, 1985 (PubMed).

Thomson, Kalishman, Golos et al.: "Isolation of a primate embryonic stem cell line." in: Proceedings of the National Academy of Sciences of the United States of America, Vol. 92, Issue 17, pp. 7844-8, 1995 (PubMed).

Thomson, Itskovitz-Eldor, Shapiro et al.: "Embryonic stem cell lines derived from human blastocysts." in: Science (New York, N.Y.), Vol. 282, Issue 5391, pp. 1145-7, 1998 (PubMed).

Badcock, Pigott, Goepel et al.: "The human embryonal carcinoma marker antigen TRA-1-60 is a sialylated keratan sulfate proteoglycan." in: Cancer research, Vol. 59, Issue 18, pp. 4715-9, 1999 (PubMed).

Draper, Pigott, Thomson et al.: "Surface antigens of human embryonic stem cells: changes upon differentiation in culture." in: Journal of anatomy, Vol. 200, Issue Pt 3, pp. 249-58, 2002 (PubMed).

Henderson, Draper, Baillie et al.: "Preimplantation human embryos and embryonic stem cells show comparable expression of stage-specific embryonic antigens." in: Stem cells (Dayton, Ohio), Vol. 20, Issue 4, pp. 329-37, 2002 (PubMed).

Schopperle, DeWolf: "The TRA-1-60 and TRA-1-81 human pluripotent stem cell markers are expressed on podocalyxin in embryonal carcinoma." in: Stem cells (Dayton, Ohio), Vol. 25, Issue 3, pp. 723-30, 2007 (PubMed).

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