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Alexa Fluor® 647 Mouse Anti-Human Terminal Transferase (TdT)

BD Pharmingen™ Alexa Fluor® 647 Mouse Anti-Human Terminal Transferase (TdT)

Clone E17-1519

(RUO)
Alexa Fluor® 647 Mouse Anti-Human Terminal Transferase (TdT)
Flow cytometric analysis of TdT expression by human REH cells. Cells from the human REH (Acute B cell leukemia, ATCC CRL-8283) cell line were fixed with 1% paraformaldehyde and permeabilized with 0.1% Triton-X100. The cells were then stained with either Alexa Fluor® 647 Mouse IgG1 κ Isotype Control (Cat. No. 557732; dashed line histogram) or Alexa Fluor® 647 Mouse Anti-Human Terminal Transferase (TdT) antibody (Cat. No. 565231; solid line histogram). The fluorescence histogram showing Terminal Transferase (TdT) expression (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of intact REH cells. flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
Flow cytometric analysis of TdT expression by human REH cells. Cells from the human REH (Acute B cell leukemia, ATCC CRL-8283) cell line were fixed with 1% paraformaldehyde and permeabilized with 0.1% Triton-X100. The cells were then stained with either Alexa Fluor® 647 Mouse IgG1 κ Isotype Control (Cat. No. 557732; dashed line histogram) or Alexa Fluor® 647 Mouse Anti-Human Terminal Transferase (TdT) antibody (Cat. No. 565231; solid line histogram). The fluorescence histogram showing Terminal Transferase (TdT) expression (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of intact REH cells. flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
Product Details
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BD Pharmingen™
TdT; DNTT; Terminal transferase; Terminal addition enzyme
Human (QC Testing)
Mouse BALB/c IgG1, κ
Purified Human TdT
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
1791
AB_2739125
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO


Preparation And Storage

Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to Alexa Fluor® 647 under optimum conditions, and unreacted Alexa Fluor® 647 was removed.

Recommended Assay Procedures

1. Harvest cultured target cells into a 50 ml conical centrifuge tube. Centrifuge at 1000 rpm for 10 minutes, aspirate and discard supernatant.

2. Wash cell pellet once with PBS and mix gently. Centrifuge at 1000 rpm for 10 minutes, aspirate and discard supernatant.

3. Fix the cells by adding 15-20 ml of 1% formaldehyde while vortexing the pellet and incubate for 20 minutes at room temperature. Centrifuge at 1000 rpm for 10 minutes, aspirate and discard supernatant.

4. Add 15-20 ml of 0.1% Triton™ X-100 in PBS and incubate for 5-10 minutes. Centrifuge at 1000 rpm for 10 minutes, aspirate and discard supernatant.

5. Resuspend cells in PBS + 1% FBS (wash buffer) to a final concentration of approximately 1 x 10^6 cells per 50 µl.

6. Prepare one tube with 50 µl of cell suspension and add 20 µl of fluorescent Anti-Human TdT antibody. Prepare another tube with 50 µl of cell suspension and add 20 µl of a matched fluorescent Ig isotype control. Mix the tube contents gently, and incubate in the dark at room temperature for 20-30 minutes.

7. Wash cells with 2 ml of wash buffer per tube. Centrifuge for 5 minutes at 1000 rpm, aspirate and discard supernatant.

8. Resuspend cells in 500 µl of wash buffer and analyze by flow cytometry.

Product Notices

  1. This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
  2. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  3. Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
  4. The Alexa Fluor®, Pacific Blue™, and Cascade Blue® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc. for research use only, excluding use in combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430), Pacific Blue™ dye, and Cascade Blue® dye are covered by pending and issued patents.
  5. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
  6. Triton is a trademark of the Dow Chemical Company.
  7. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  8. 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.
  9. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  10. An isotype control should be used at the same concentration as the antibody of interest.
565231 Rev. 1
Antibody Details
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E17-1519

The E17-1519 monoclonal antibody specifically recognizes Terminal deoxynucleotidyl transferase (TdT), which is also known as Terminal transferase, or DNA nucleotidylexotransferase (DNTT). TdT is a 60 kDa nuclear polymerase that catalyzes the template-independent addition of deoxynucleotides to the 3'-hydroxyl termini of single-stranded oligonucleotide primer segments. TdT can contribute to antigen receptor diversity by N region diversification of rearranging immunoglobulin heavy chain and T cell receptor gene segments during pre-B and pre-T cell development. It is also expressed by certain lymphomas and leukemias.

        

565231 Rev. 1
Format Details
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Alexa Fluor™ 647
Alexa Fluor™ 647 Dye is part of the BD red family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 653-nm and an emission maximum (Em Max) at 669-nm. Alexa Fluor 647 is designed to be excited by the Red laser (627-640 nm) and detected using an optical filter centered near 520-nm (e.g., a 660/20 nm bandpass filter). Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
Alexa Fluor™ 647
Red 627-640 nm
653 nm
669 nm
565231 Rev.1
Citations & References
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View product citations for antibody "565231" on CiteAb

Development References (4)

  1. Fuller SA, Philips A, Coleman MS. Affinity purification and refined structural characterization of terminal deoxynucleotidyltransferase. Biochem J. 1985; 231(1):105-113. (Biology). View Reference
  2. Gopcsa L, Banyai A, Jakab K, et al. Extensive flow cytometric characterization of plasmacytoid dendritic cell leukemia cells.. Eur J Haematol. 2005; 75(4):346-51. (Clone-specific: Flow cytometry). View Reference
  3. Keren DF, Hanson CA, Hurtubise PE. David F. Keren, Curtis A. Hanson, Paul E. Hurtubise., ed. Flow cytometry and clinical diagnosis. Chicago: ASCP Press; 1994:1-676.
  4. Sasaki R, Yuasa Y, Masuyama A, Takaku F, Bollum FJ. Production of a specific monoclonal antibody to terminal deoxynucleotidyl transferase (TdT) and the extensive studies of TdT in patients with hematological malignancies. 1993; 25(4):223-225. (Biology). View Reference
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565231 Rev. 1

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Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

For Research Use Only. Not for use in diagnostic or therapeutic procedures.