| Description | The HA tag consists of the amino acid sequence YPYDVPDYA (residues 98-106 of human influenza hemagglutinin). It has minimal impact on the tertiary structure of the target foreign protein and can be easily fused to either the N- or C-terminus, making it a popular choice for recombinant protein The HA tag consists of the amino acid sequence YPYDVPDYA (residues 98-106 of human influenza hemagglutinin). It has minimal impact on the tertiary structure of the target foreign protein and can be easily fused to either the N- or C-terminus, making it a popular choice for recombinant protein expression. Anti-HA Agarose Resin is based on a 4% agarose gel matrix, which minimizes non-specific binding of host cell proteins, making it suitable for both the purification and immunoprecipitation (IP) of HA-tagged fusion proteins.Aladdin Anti-HA Agarose Resin is stored in a solution containing 0.1% ProClin 300, with a settled gel to storage solution ratio of 1:1. The product specification refers to the actual volume of the settled gel.ParameterValue / DescriptionMatrix4% Agarose MicrospheresLigandAnti-HA Mouse Monoclonal AntibodyParticle Size Range45~165 µmBinding Capacity>1 mg HA-tagged protein / mL resinMaximum Pressure0.1 MPa, 1 barStorage Conditions0.1% ProClin 300, 2~8℃Shelf Life2 yearsProtocol1. Sample PreparationEnsure the sample solution has appropriate ionic strength and pH before loading. Dilute the sample or cell culture supernatant with equilibration buffer, or dialyze the sample against equilibration buffer.Clarify the sample by centrifugation or filtration through a 0.22 µm or 0.45 µm membrane to reduce impurities, improve purification efficiency, and prevent column clogging.2. Buffer PreparationIt is recommended to filter water and buffers through a 0.22 µm or 0.45 µm membrane before use.Equilibration Buffer: 10 mM Tris, 0.15 M NaCl, pH 7.4Wash Buffer: 10 mM Tris, 0.15 M NaCl, 0.05% Tween-20, pH 7.4Chemical Elution Buffers:0.1 M Glycine-HCl, pH 2.0–2.83 M Sodium Thiocyanate (NaSCN)50 mM NaOHCompetitive Elution Buffer: 50 mM Tris, 0.15 M NaCl, 100–500 µg HA peptide / mL, pH 7.4Neutralization Buffer: 1 M Tris-HCl, pH 8.5Comparison of Chemical Elution BuffersSolutionAdvantagesDisadvantages0.1M glycine HCl, pH 2.0-2.8Does not damage resin binding capacity if target protein is stable at low pHLow elution efficiency; target protein may denature3M NaSCNHigh elution efficiency; does not damage resin binding capacityTarget protein may denature50mM NaOHHigh elution efficiencyTarget protein may denature; reduces resin lifespan3. Sample Purification3.1 Column Chromatography(1) Pack the Anti-HA Agarose Resin into a suitable chromatography column. Equilibrate the column with 5 column volumes (CV) of Equilibration Buffer.(2) Load the sample onto the equilibrated resin. Collect the flow-through. The sample can be reloaded to increase binding efficiency.(3) Wash with 10–20 CV of Wash Buffer to remove non-specifically bound proteins. Collect the wash fractions.(4) Elution:* A. Acidic Elution: Elute with 5 CV of acidic elution buffer (e.g., 0.1 M Glycine-HCl, pH 2.0–2.8). Add a volume of Neutralization Buffer equal to one-tenth of the elution volume to each fraction to adjust the pH to 7.0–8.0. Collect fractions separately.* Note: After acidic elution, the resin must be immediately re-equilibrated. Do not expose the resin to the acidic elution buffer for more than 20 minutes.* B. Chemical Elution: Elute with 5 CV of a chemical elution buffer (e.g., 3 M NaSCN or 50 mM NaOH). Collect fractions separately.* Note: After chemical elution, the resin must be immediately re-equilibrated. Do not expose the resin to the chemical elution buffer for more than 20 minutes.* C. Competitive Elution: Elute with 5 CV of Competitive Elution Buffer. Collect fractions separately.(5) Regenerate the resin with 3 CV of a chemical elution buffer (e.g., Glycine-HCl), then re-equilibrate with Equilibration Buffer until neutral pH is reached.(6) Store the resin in Storage Buffer at 2–8°C.3.2 Batch/Binding Method(1) Resin Preparation: Transfer an appropriate amount of Anti-HA Agarose Resin to a column and drain the storage solution. Wash with 5 CV of Equilibration Buffer.(2) Add the sample solution. Incubate with shaking at 4°C or room temperature for 30 minutes (avoid magnetic stirring). Ensure thorough mixing.(3) After incubation, centrifuge the mixture (5,000 × g, 1 min) or filter to collect the resin.(4) Transfer the resin to a column. Wash with Equilibration Buffer until the UV baseline stabilizes.(5) Elute using either the Chemical or Competitive Elution method as described in section 3.1 (4).(6) Regenerate and store the resin as described in sections 3.1 (5) and (6).3.3 Immunoprecipitation (IP) Procedure(1) Resin Preparation: Add 40 µL of Anti-HA Agarose Resin suspension (20 µL settled resin) to a 1.5 mL tube. Centrifuge at 5,000 × g for 1 min. Discard the supernatant.(2) Add 0.5 mL of Equilibration Buffer to resuspend the resin. Centrifuge at 5,000 × g for 1 min. Discard the supernatant. Repeat this wash step once.(3) Add 200–1000 µL of sample lysate to the prepared resin. Mix and incubate on a tube rotator at room temperature for at least 1 hour. Centrifuge at 5,000 × g for 1 min. Collect the supernatant for analysis.(4) Washing: Add 0.5 mL of Wash Buffer, resuspend the resin, and mix gently. Centrifuge at 5,000 × g for 1 min. Discard the supernatant. Repeat this wash step three more times.(5) Elution: Choose the elution method based on downstream application.* A. Chemical Elution: Add 100 µL of chemical elution buffer (0.1 M Glycine-HCl pH 2.0-2.8, 3 M NaSCN, or 50 mM NaOH) and resuspend the resin. Incubate at room temperature for 5 min. Centrifuge at 5,000 × g for 1 min. Carefully collect the supernatant and neutralize immediately if acidic. Store eluted samples at 4°C short-term or -20°C long-term.* B. Competitive Elution: Add 100 µL of Competitive Elution Buffer and resuspend the resin. Incubate at room temperature for 30 min. Centrifuge at 5,000 × g for 1 min. Carefully collect the supernatant. Repeat elution 1-2 times. Store eluted samples at 4°C short-term or -20°C long-term.* C. Denaturing Elution (SDS-PAGE): Add 20 µL of 2× Loading Buffer (contains SDS and reducing agents like β-mercaptoethanol/DTT) to the resin. Heat at 95°C for 5 min. Centrifuge at 5,000 × g for 1 min, and load the supernatant directly onto an SDS-PAGE gel for analysis. Note: This method denatures the antibody, rendering the resin unusable for reuse.Troubleshooting Guide... Read More | Inquire | Product IntroductionHuman epidermal growth factor (EGF) has a molecular weight of 6 KD, contains 53 amino acids, and has three intramolecular disulfide bonds.Upon binding to EGFR, a specific receptor located on the cell surface, EGF causes a series of biochemical reactions in the cell: increasing Product IntroductionHuman epidermal growth factor (EGF) has a molecular weight of 6 KD, contains 53 amino acids, and has three intramolecular disulfide bonds.Upon binding to EGFR, a specific receptor located on the cell surface, EGF causes a series of biochemical reactions in the cell: increasing the level of calcium ion concentration in the cell, promoting the progress of glycolysis, increasing the synthesis of proteins, and can enhance the expression of EGFR, a specific class of genes to promote DNA synthesis and cell proliferation.Specification parametersSource Pichia pastorisAppearance white lyophilized powderActivity ≥1.0×106IU/mgpH 6.5-7.5Molecular weight 6.5kDEndotoxin ≦1.0 EU/mgCAS No 62253-63-8Matters needing attentionReconstitution: reconstitution of REGF lyophilized powder to 100-200 µg/ml with sterile water is recommended and further dilution with other solvents may be performed.REGF dissolved at 4 ° C can be stored for 2-7 days and used up as soon as possible.To not use for short periods, store at - 20 ℃.Use as soon as possible after opening to avoid contamination.Limitations of useIt is suitable for research, laboratory and production use only and cannot be used directly in humans... Read More | Purity:>95%, by SDS-PAGE visualized with Coomassie® Blue Staining. Description： Interleukin 33 (IL-33), also known as DVS27 or NF-HEV (Nuclear Factor from High Endothelial Venules), is a pro-inflammatory protein and a chromatin-associated cytokine of the IL-1 family with high sequencePurity:>95%, by SDS-PAGE visualized with Coomassie® Blue Staining. Description： Interleukin 33 (IL-33), also known as DVS27 or NF-HEV (Nuclear Factor from High Endothelial Venules), is a pro-inflammatory protein and a chromatin-associated cytokine of the IL-1 family with high sequence and structural similarity to IL-1 and IL-18. IL-33 protein is expressed highly and rather selectively by high endothelial venule endothelial cells (HEVECs) in human tonsils, Peyer's patches, and lymph nodes. IL-33 protein has transcriptional regulatory properties, and the researches suggested that IL-33 is a dual-function protein that might act both as a cytokine and as an intracellular nuclear factor. As a type 2 cytokines, IL-33 protein also play a pivotal role in helminthic infection and allergic disorders... Read More | Fibroblast growth factor-21 (FGF-21) belongs to the large FGF family and it is specifically induced by HMGCS2 activity. In mice, brown adipose tissue becomes a source of systemic FGF21 after cold exposure. FGF-21 stimulates glucose uptake in differentiated adipocytes via the induction of glucose Fibroblast growth factor-21 (FGF-21) belongs to the large FGF family and it is specifically induced by HMGCS2 activity. In mice, brown adipose tissue becomes a source of systemic FGF21 after cold exposure. FGF-21 stimulates glucose uptake in differentiated adipocytes via the induction of glucose transporter SLC2A1/GLUT1 expression and the activity depends on the presence of KLB. FGF-21, in the presence of β-Klotho as a protein cofactor, signals through the FGFR 1c and 4 receptors. Murine FGF-21 shows limited binding to heparin. In addition, Murine FGF-21 respectively shows 81% and 92% a.a. identity to human and rat FGF-21, and it show activity on human and rat cells. Recombinant Murine FGF21 is a 19.9kDa globular protein containing 182 amino acid residues.Purity>96%(SDS-PAGE, HPLC)Additional sequence informationA single non-glycosylated polypeptide chain containing 182 amino acids. This product is for the mature full length protein. The signal peptide is not included.FunctionStimulates glucose uptake in differentiated adipocytes via the induction of glucose transporter SLC2A1/GLUT1 expression (but not SLC2A4/GLUT4 expression). Activity requires the presence of KLB... Read More |