Human Transferrin

• Cat.No.: TF-4777H
• Product Overview: Product is the lyophilized powder of human TF and buffer salts. Human TF is purified from pooled human serum using multi-step procedures which may include salt fractionation, gel filtration, ion-exchange chromatography and immunoabsorption. The product is dialyzed into 0.01M sodium phosphate, 0.07M sodium chloride, pH 7.3, filtered through a 0.22 µm filter, vialed and lyophilized. No preservative has been added.

Specification

• Species: Human
• Source: Human Plasma
• Tag: Non
• Description: TF is a glycoprotein that binds iron very tightly but reversibly. Although iron bound to transferrin is less than 0.1% (4 mg) of the total body iron, it is the most important iron pool, with the highest rate of turnover (25 mg/24 h). TF has a molecular weight of around 80 kDa and contains 2 specific high-affinity Fe (III) binding sites.
• Preparation: The protein is purified from pooled human plasma and lyophilized from 0.02M NH4HCO3.
• Reconstitution: The protein is readily soluble at 1% in deionised. Alternatively, it can be reconstituted with a neutral buffer, such as PBS.
• SDS-PAGE: The preparation shows a tight cluster of bands with a molecular weight of 77 kDa. Dimers may appear under certan conditions.
• Protein: 98% (By Lowry)
• Purity: > 98 % (By Cellulose Acetate Electrophoresis)
• Iron: 27.85 µg/g (By Atomic Absorption). 1 gm transferrin can bind approximately 1400-1500 µg of iron.
• pH: 7.24 (3% solution)
• Endotoxin Level: 0.034 EU/mg (By Limulus Amebocyte Lysate)
• Microbial detection: No Mycoplasma is detected and Bioburden is less than 10 cfu/ml.
• Moisture: 0.9 % (By Loss on Drying)
• Storage: Store at -10°C or below.
• Pathways: EPHB forward signaling; Formation of Platelet plug; HIF-1-alpha transcription factor network; Hemostasis; Iron uptake and transport; Mineral absorption; Platelet Activation; Platelet degranulation; Response to elevated platelet cytosolic Ca2+; Transferrin endocytosis and recycling; Transmembrane transport of small molecules

Gene Information

• Gene Name: TF transferrin [ Homo sapiens ]
• Official Symbol: TF
• Synonyms: TF; transferring; PRO1557; PRO2086; DKFZp781D0156; serotransferrin; siderophilin; OTTHUMP00000197155; beta-1 metal-binding globulin; Serotransferrin; Beta-1 metal-binding globulin; Siderophilin
• Gene ID: 7018
• mRNA Refseq: NM_001063
• Protein Refseq: NP_001054
• MIM: 190000
• UniProt ID: P02787
• Chromosome Location: 3q21
• Function: ferric iron binding; metal ion binding; protein binding; ubiquitin protein ligase binding

Citation

Morphological cell profiling of SARS-CoV-2 infection identifies drug repurposing candidates for COVID-19

Journal: Proceedings of the National Academy of Sciences of the United States of America    PubMed ID: 34413211    Data: 2021/9/7

Authors: Carmen Mirabelli, Jesse W. Wotring, Jonathan Z. Sexton

Article Snippet:The library was formatted in five 384-well compound plates and was dissolved in DMSO at 10 mM.The library was formatted in five 384-well compound plates and was dissolved in DMSO at 10 mM.. Apolactoferrin was provided by Glanbia Nutritionals (Bioferrin 2000); Hololactoferrin (Sigma-Aldrich; L4765), native human lactoferrin (Creative BioMart; LFT-8196H), and transferrin (Sigma-Aldrich; T2036) were handled separately and added manually in cell culture media.. Dilution plates were generated for qHTS at concentrations of 2 mM, 1 mM, 500 μM, 250 μM, and 50 μM, and compounds were dispensed at 1:1,000 dilution.Dilution plates were generated for qHTS at concentrations of 2 mM, 1 mM, 500 μM, 250 μM, and 50 μM, and compounds were dispensed at 1:1,000 dilution.

Lactoferrin blocks SARS-CoV-2 at the attachment step. ( A ) Huh7 cells were infected with SARS-CoV-2 at MOI of 0.2 for 48 h and treated with increasing concentration of lactoferrin (6.25–6,250 nM). Cells were harvested and RNA was extracted. Viral genome copies were calculated by RT-qPCR with an absolute quantification method. ( B ) Huh7 were infected with SARS-CoV-2 (MOI of 1, 5, and 10; MOI of 0 indicates noninfected cells) and treated with 6,250 nM lactoferrin at 1 and 24 h p.i. Bars indicate the percentage of infected cells in different conditions. Data are an average of eight replicates. Statistical significance determined using multiple Student’s t test with the Bonferroni–Dunn method, with α = 0.05. Except for MOI of 0, all conditions (untreated vs. lactoferrin, 1 h or untreated vs. lactoferrin, 24 h) differ by P < 0.0001. ( C ) Percentage of SARS-CoV-2–infected Huh7 cells upon treatment with bovine apolactoferrin and hololactoferrin, native human lactoferrin, and transferrin at a concentration of 6,250 nM. ( D ) Binding assay. Huh7 cells were preincubated on ice with compounds: lactoferrin (1,250 and 6,250 nM) and remdesivir (10 nM) as negative controls for 1 h and then infected with SARS-CoV-2 (MOI = 10) for 1 h on ice. Cells were then washed thoroughly with PBS to remove unbound virus and viral RNA was quantified by RT-qPCR. Huh7 were cultured in NaClO 3 for 7 d, which strips heparan sulfate proteoglycans from the cell surface, and were subsequently used as a control for lactoferrin mode of action. ( E ) Synergy analysis of lactoferrin in combination with remdesivir. Cells were pretreated with combinations or single agents and infected with SARS-CoV-2 (MOI of 10) for 48 h. Data are shown after normalization to viral control (100%) and represent an average of n = 3 biological replicates with n = 2/3 technical replicates each. Unpaired t tests with Welch’s correction were performed in GraphPad Prism to determine significance. * P < 0.0001.

Lactoferrin indirect antiviral action is mediated by the up-regulation of cellular innate immune response. iAEC2 cells were treated with lactoferrin (6,250 nM) for 48 h, and then RNA was extracted and sequenced. ( A ) Volcano plot of genes down-regulated (blue) and up-regulated (red) upon treatment with lactoferrin. ( B ) Analysis of the top up-regulated and down-regulated pathways. ( C ) Heatmap of selected genes. In red are highlighted genes associated with inflammation and antiviral response.