Active Recombinant Human JAG1 protein, Fc-tagged

• Cat.No.: JAG1-3138H
• Product Overview: Active Recombinant Human JAG1 protein(NP_000205.1)(Met 1-Ser 1046) was expressed in HEK293, fused with Fc region of human IgG1 at the C-terminus.

Specification

• Species: Human
• Source: HEK293
• Tag: Fc
• Protein Length: 1-1046 aa
• Form: Lyophilized from sterile PBS, pH 7.4.
• Bio-activity: Measured by the ability of the immobilized protein to enhance BMP2 induced alkaline phosphatase activity in C3H10T1/2 mouse embryonic fibroblast cells. The ED50 for this effect is typically 5-30 μg/mL.
• Molecular Mass: The recombinant human JAG1/Fc is a disulfide-linked homodimer. The reduced monomer consists of 1254 amino acids and has a predicted molecular mass of 137 kDa. In SDS-PAGE under reducing conditions, the apparent molecular mass of rh JAG1/Fc monomer is approximately 180-200 kDa due to glycosylation.
• Endotoxin: < 1.0 EU per μg of the protein as determined by the LAL method.
• Purity: > 85 % as determined by SDS-PAGE
• Storage: Store it under sterile conditions at -20°C to -80°C upon receiving. Recommend to aliquot the protein into smaller quantities for optimal storage. Avoid repeated freeze-thaw cycles.
• Reconstitution: It is recommended that sterile water be added to the vial to prepare a stock solution of 0.2 ug/ul. Centrifuge the vial at 4°C before opening to recover the entire contents.
• Publications:
  • A non-canonical JAGGED1 signal to JAK2 mediates osteoblast commitment in cranial neural crest cells (2019)
  • The Special Role of JAK/STAT, and Notch Signaling Pathways in Cancer Pathogenesis (2023)
  • Delivery of A Jagged1-PEG-MAL hydrogel with Pediatric Human Bone Cells Regenerates Critically-Sized Craniofacial Bone Defects (2023)
  • Controlled JAGGED1 delivery induces human embryonic palate mesenchymal cells to form osteoblasts (2017)
  • JAGGED1 stimulates cranial neural crest cell osteoblast commitment pathways and bone regeneration independent of canonical NOTCH signaling (2021)
  • Osteoinductive effect of soluble transforming growth factor beta receptor 3 on human osteoblast lineage (2021)

Gene Information

• Gene Name: JAG1 jagged 1 [ Homo sapiens ]
• Official Symbol: JAG1
• Synonyms: JAG1; jagged 1; AGS, Alagille syndrome, JAGL1; protein jagged-1; AHD; AWS; CD339; HJ1; AGS; JAGL1; MGC104644
• Gene ID: 182
• mRNA Refseq: NM_000214
• Protein Refseq: NP_000205
• UniProt ID: P78504

Citation

JAGGED1 Stimulates Cranial Neural Crest Cell Osteoblast Commitment Pathways and Bone Regeneration Independent of Canonical NOTCH Signaling

Journal: Bone    PubMed ID: 32980561    Data: 2022/4/24

Authors: Archana Kamalakar, Jay M. McKinney, Steven L. Goudy

Article Snippet:50 μL (1.5 mg) of Dynabeads Protein G ( 36 ) (Invitrogen 10003D) were transferred to a tube, where the beads were separated from the solution using a magnet.50 μL (1.5 mg) of Dynabeads Protein G ( 36 ) (Invitrogen 10003D) were transferred to a tube, where the beads were separated from the solution using a magnet.. Recombinant JAG1-Fc (5 μM, 5.7 μM, 10 μM or 20 μM) (Creative BioMart, JAG1–3138H) and IgG-Fc fragment (5 μM or 5.7 μM) (Abcam, ab90285) alone were diluted in 200 μL PBS with 0.1% Tween-20 (Fisher, BP337–500) and then added to the dynabeads.. The beads plus proteins were incubated at 4°C with rotation for 16 hours.The beads plus proteins were incubated at 4°C with rotation for 16 hours.

As a proof of concept experiment, we (A) incorporated JAG1-dynabeads complex (5 μM, 10 μM or 20 μM), dynabeads alone and BMP2 (2.5 μM) in 4% PEG-MAL hydrogels and implanted them into (B-G) 3.5 mm critical-sized defects in the parietal bones of 6–8-week old C57BL/6 mice and delivered them (J) without or (K) with CNC cells into the defects as 3 separate doses (Initial dose, Week 4, Week 8), as shown in H. After 12 weeks, we quantified differences in regenerated bone volume (J-K) within the defect and compared them between experimental groups by μCT analysis. μCT reconstructions of defects are shown in I. Data were subjected to ANOVA and Tukey’s post-test and are presented as mean (n ≥ 2) ± SD with p values reported.

(A) A principal component analysis of all treatments revealed that PC1 distinguished JAG1-treated cells to the right from non-JAG1 treated cells to the left along the horizontal axis. PC2 distinguished cells treated with JAG1 alone towards the top and JAG1 + DAPT treated cells towards the bottom along the vertical axis. (B) Clustering analysis using Pearson correlation-coefficient reveals the heatmap showing distinct gene clusters downstream of different treatments (n = 3). Cluster A and B consists of genes downregulated by JAG1 and Cluster C consists of genes upregulated by JAG1.

(A-B) Volcano plots of differentially expressed genes in CNC cells that were treated with JAG1-dynabeads complex (5.7 μM) with or without DAPT (15 μM) compared to Fc-dynabeads complex (5.7 μM). (C) Subsequent comparison of the JAG1 and JAG1 + DAPT treatment groups for genes that were significantly upregulated in either group compared to Fc. Plots indicate genes that were down-regulated in response to DAPT (red) or preserved between groups (grey). (D) Comparison between JAG1 treated-samples in the presence and absence of DAPT revealed various distinct genes, like Cxcl1, Cxcl12, Hes1 and Il6 upregulated downstream of JAG1 alone and Prl2c2 upregulated downstream of JAG1 + DAPT along with some conserved genes like Id1 and Rhou. Data were subjected to ANOVA and Tukey’s post-test and are presented as mean (n ≥ 2) ± SD with p values reported.

Controlled JAGGED1 delivery induces human embryonic palate mesenchymal cells to form osteoblasts

Journal: Journal of biomedical materials research. Part A    PubMed ID: 28913955    Data: 2019/2/1

Authors: Jean De La Croix Ndong, Yvonne Stephenson, Steven Goudy

Article Snippet:Culture well chambered coverglass were pre-coated with rabbit anti-human IgG (10 μg/mL) in phosphate saline buffer (PBS) for 30 min at 37°C and subsequently blocked with cell culture growth medium for 30 min. Chambered coverglass were then coated with 5 μg/mL JAGGED1/Fc (JAG1–3138 H, Creative BioMart) diluted in growth media for 2 hours at 37°C.. As control for JAGGED1, human IgG (5 μg/mL) was used.As control for JAGGED1, human IgG (5 μg/mL) was used.

JAGGED1 release profile and Notch signaling activation. (A) Prefunctionalized PEG-MAL with JAGGED1 (10 ug/m) and RGD peptide were cross-linked and incubated for 20 days in PBS. Released JAGGED1 from PEG-MAL collected every 2–3 days and quantified by ELISA. (B–C). HEPM cells were encapsulated in functionalized JAGGED1–5%PEG-MAL hydrogels and cultured in growth media for 3 days. HEPM cells proliferation (B) and Apoptosis were measured using the CellTiter Aqueous One 96 Kit and Apo-One Homogeneous Caspase3/7 kit, respectively. (n = 3) *p <0.05 versus IgG control (JAGGED1 (0 ug/mL) and #p <0.05 versus day 0.

Activation of Notch Signaling Pathway by Jagged-1PEG-MAL. HEPM cells or Notch reporter CHO cells were encapsulated in functionalized JAGGED1–5%PEG-MAL hydrogels and cultured for 3 days. (A) Notch signaling activation in CHO cells analyzed by YFP fluorescence and quantified using Image J. (bar: 100 μm). (B) Notch-associated gene expression in HEPM cells were assessed by RT-PCR. (n = 3) *p <0.05 versus IgG control (JAGGED1 (0 μg/mL).

JAGGED1-PEG-MAL stimulates the differentiation of HEPM cells in vitro. HEPM cells were encapsulated in functionalized JAGGED1–5%PEG-MAL hydrogels and cultured in osteogenic media for 7 days. (A) HEPM differentiation analyzed by ALP activity using p-nitrophenyl phosphate (pNPP) as a substrate which turns yellow when dephosphorylated by ALP. (B) Expression of osteoblast marker genes was assessed by qPCR (n = 3) *p <0.05 versus IgG control (JAGGED1 (0 μg/mL).

Reviews

07/31/2024

JAG1-3138H

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