Recombinant Human Flt-3 Ligand/FLT3L GMP Protein, CF

GMP-grade Recombinant Human Flt-3 Ligand/FLT3L Protein (Catalog # BT-FT3L-GMP) as measured in a cell proliferation assay using OCIAML5 acute myeloid leukemiacells. Three independent lots were tested for activity and plotted on the same graph to show lot-to-lot consistency of GMP Flt-3 Ligand/FLT3L.

Equivalent bioactivity of GMP (Catalog # BT-FT3L-GMP) and Animal-Free (BT-FT3L-AFL) grades of Recombinant Human Flt-3 Ligand/FLT3L as measured in a cell proliferation assay using OCI-AML5 acute myeloid leukemia cells (orange and green, respectively).

2 μg/lane of Recombinant Human Flt-3 Ligand/FLT3L GMP Protein (Catalog # BT-FT3L-GMP) was resolved with SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by Coomassie® Blue staining, showing bands at 17 kDa, under reducing conditions.

Purified CD34+ cells derived from plerixafor mobilized peripheral blood were cultured in commercially available media. The media was supplemented with SCF (100 ng/mL, Catalog # BT-SCF-GMP), TPO (100 ng/mL, Catalog # BT-TPO-GMP), FLT-3L (100 ng/mL, Catalog # BT-FT3L-GMP), and SR1 (1 µM, Catalog # 7086). After 7 days, fold expansion was calculated from total viable cell counts. Frequency (%) of CD34+ cells was analyzed by flow cytometry on Days 0 and 7. The average fold expansion of n=2 technical replicates is shown.

Purified CD34+ cells derived from plerixafor mobilized peripheral blood were cultured in commercially available media. The media was supplemented with SCF (Catalog # BT-SCF-GMP, 100 ng/mL), TPO (Catalog # BT-TPO-GMP, 100 ng/mL), FLT-3L (Catalog # BT-FT3L-GMP, 100 ng/mL), and SR1 (Catalog # 7086, 1 µM). After 7 days, fold expansion was calculated from total viable cell counts. Percent (%) CD34+ cells was analyzed by flow cytometry on Days 0 and 7. The average fold expansion of n=3 technical replicates is shown.

• Reactivity: Hu
• Applications: Bioactivity
• Format: Carrier-Free

Recombinant Human Flt-3 Ligand/FLT3L GMP Protein, CF Summary

• Details of Functionality: Measured in a cell proliferation assay using OCI-AML5 acute myeloid leukemia cells. The ED₅₀ for this effect is 0.200-2.00 ng/mL.
  The specific activity of Recombinant Human Flt-3 Ligand/FLT3L is >5.00 x 10⁵ units/mg, which is calibrated against the human Flt-3 Ligand WHO standard (NIBSC code: 96/532).
• Source: E. coli-derived human Flt-3 Ligand/FLT3L protein
  Thr27 - Ala181
  Produced using non-animal reagents in an animal-free laboratory.
  Manufactured and tested under cGMP guidelines.
• Accession #: P49771.1
• N-terminal Sequence: Met-Thr27-Gln-Asp-(Cys)-Ser-Phe-Gln-His-Ser & Thr27-Gln-Asp-(Cys)-Ser-Phe-Gln-His-Ser-Pro
• Protein/Peptide Type: GMP Recombinant Proteins
• Purity: >97%, by SDS-PAGE with quantitative densitometry by Coomassie® Blue Staining. The molecular weight by mass spectrometry is 17731 Da ± 50 Da, with a second mass of 17600 Da ± 50 Da also present
• Endotoxin Note: <0.10 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

• Dilutions:
  • Bioactivity
• Theoretical MW: 18 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
• SDS-PAGE: 17 kDa, under reducing conditions.

Packaging, Storage & Formulations

• Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • A minimum of 12 months when stored at ≤ -20 °C as supplied. Refer to lot specific COA for the Use by Date.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 3 months, ≤ -20 °C under sterile conditions after reconstitution.
• Buffer: Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose.
• Purity: >97%, by SDS-PAGE with quantitative densitometry by Coomassie® Blue Staining. The molecular weight by mass spectrometry is 17731 Da ± 50 Da, with a second mass of 17600 Da ± 50 Da also present
• Reconstitution Instructions: Reconstitute at 500 μg/mL in PBS.

Notes

Full terms and conditions of sale can be found online in the Protein Sciences Segment T&Cs at: Terms & Conditions.

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Human Flt-3 Ligand/FLT3L GMP Protein, CF

• FL
• FLG3L
• Flt3 ligand
• Flt-3 Ligand
• Flt3L
• FLT3LG
• fms-related tyrosine kinase 3 ligand
• SL cytokine

Background

Flt‑3 Ligand, also known as FLT3L, is an alpha-helical cytokine that promotes the differentiation of multiple hematopoietic cell lineages (1-3). Mature human Flt‑3 Ligand consists of a 158 amino acid (aa) extracellular domain (ECD) with a cytokine-like domain and a juxtamembrane tether region, a 21 aa transmembrane segment, and a 30 aa cytoplasmic tail (4-7). Within the ECD, human Flt‑3 Ligand shares 71% and 65% aa sequence identity with mouse and rat Flt‑3 Ligand, respectively (4-6). The human and mouse Flt‑3 Ligand proteins show cross-species activity. Flt-3 Ligand is also structurally related to M-CSF and SCF. Flt-3 Ligand is widely expressed in various human and mouse tissues. It is expressed as a noncovalently-linked dimer by T cells and bone marrow and thymic fibroblasts (1, 8). Each 36 kDa chain of the Flt-3 Ligand dimer carries approximately 12 kDa of N- and O-linked carbohydrates (8). Alternate splicing and proteolytic cleavage of the transmembrane form of the Flt-3 Ligand protein can generate a soluble 30 kDa fragment that includes the cytokine-like domain (4, 8). Alternate splicing of human Flt‑3 Ligand also generates membrane-associated isoforms that contain either a truncated cytoplasmic tail or an 85 aa substitution following the cytokine-like domain in the ECD of the Flt-3 Ligand protein (4, 5, 8). Both transmembrane and soluble forms of Flt‑3 Ligand signal through the tyrosine kinase receptor Flt-3/Flk-2 (3, 4, 6, 7). Flt‑3 Ligand induces the expansion of monocytes and immature dendritic cells as well as early B cell lineage differentiation (2, 9). Additionally, Flt-3 Ligand synergizes with IL-3, GM-CSF, and SCF to promote the mobilization and myeloid differentiation of hematopoietic stem cells (4-6). Flt-3 Ligand also cooperates with IL-2, IL-6, IL-7, and IL-15 to induce NK cell development and with IL-3, IL-7, and IL-11 to induce terminal B cell maturation (1, 10). Animal studies show that Flt‑3 Ligand reduces the severity of experimentally induced allergic inflammation (11).

1. Wodnar-Filipowicz, A. (2003) News Physiol. Sci. 18:247.
2. Dong, J. et al. (2002) Cancer Biol. Ther. 1:486.
3. Gilliland, D.G. and J.D. Griffin (2002) Blood 100:1532.
4. Hannum, C. et al. (1994) Nature 368:643.
5. Lyman, S.D. et al. (1994) Blood 83:2795.
6. Lyman, S.D. et al. (1993) Cell 75:1157.
7. Savvides, S.N. et al. (2000) Nat. Struct. Biol. 7:486.
8. McClanahan, T. et al. (1996) Blood 88:3371.
9. Diener, K.R. et al. (2008) Exp. Hematol. 36:51.
10. Farag, S.S. and M.A. Caligiuri (2006) Blood Rev. 20:123.
11. Edwan, J.H. et al. (2004) J. Immunol. 172:5016.

Bioinformatics

• Uniprot:
  • P49771.1()