UltraBio™ NHS Magnetic Beads (1μm) - BioReagent, 10 mg/mL

Cat. No.: N1456038

GRADE & PURITY BioReagent, 10 mg/mL

Storage

Store at -20°C

Shipped In

Ice chest + Ice pads

Application

Nucleic Acid Extraction and Purification, Protein Separation and Purification

Descripción general

UltraBio™ NHS Magnetic Beads (1 μm) are pre-activated polymer magnetic microspheres designed for direct coupling with proteins or peptides containing amino groups. This pre-activated medium can be tailored to prepare specialized affinity media, enabling rapid and efficient one-step purification of target substances from complex systems. 

Parameter	Specification
Matrix	Polymer Magnetic Microspheres
Ligand	NHS
Bead Concentration	10 mg/mL
Particle Size	1 μm
Coupling Capacity	>10μg IgG/mg beads
Storage	-20°C (in 100% Isopropanol)

Instructions for Use:

I. Buffer Preparation

• Filter all water and buffers through a 0.22 μm or 0.45 μm membrane before use.

• Wash Solution: 1 mM HCl

• Coupling Buffer: 0.2 M NaHCO₃, 0.5 M NaCl, pH 8.0

• Blocking Solution: 0.5 M Ethanolamine, 0.5 M NaCl, pH 8.3 OR 0.1 M Tris, pH 8.5

• Wash Buffer 1: 0.1 M Sodium Acetate-Acetic Acid, 0.5 M NaCl, pH 3.0

• Wash Buffer 2: 0.1 M Tris-HCl, 0.5 M NaCl, pH 8.0

• Storage Solution: PBS, 0.01% Tween-20, 0.02% NaN₃

• Note: Coupling buffer can be carbonate, phosphate, or other buffers lacking primary amines. Adding salt ions reduces non-specific adsorption.

II. Sample Preparation

• Dissolve the sample in Coupling Buffer to a concentration of approximately 5-10 mg/mL.

III. Antigen Coupling

1. Take the desired amount of UltraBio™ NHS Magnetic Beads (1 μm). Aspirate the storage solution. Wash once with 1 mM HCl Wash Solution, then once with Coupling Buffer.

   • *Note: After adding wash solution and mixing, immediately place the tube on the magnetic rack for separation (~1 min), then aspirate the supernatant. Use pre-chilled solutions for rapid washing to minimize hydrolysis of the pre-activated medium.*

2. Add the dissolved sample to the washed UltraBio™ NHS Magnetic Beads (1 μm).

3. React with end-over-end rotation at 28°C for 2-4 hours or 4°C overnight.

   • Note: Ensure the beads remain suspended; settling significantly reduces coupling efficiency.

4. After reaction, collect the coupling supernatant to assess coupling efficiency. Wash the beads with deionized water. Add twice the initial bead suspension volume of Blocking Solution. React with end-over-end rotation at 28°C for 1 hour.

   • Note: UV absorbance measurement of supernatant protein concentration is unreliable post-coupling. Use SDS-PAGE or BCA assay to assess coupling efficiency.

5. Aspirate the blocking solution. Wash the beads sequentially with:

   • 3x suspension volume Deionized Water

   • 1x suspension volume Wash Buffer 1

   • 1x suspension volume Deionized Water

   • 1x suspension volume Wash Buffer 2

   • 1x suspension volume Deionized Water
     Repeat this alternating wash cycle once more (2 cycles total). Resuspend the coupled beads in Storage Solution at a concentration of 10 mg/mL. Store at 2-8°C.

IV. Purification Procedure (Example: Antigen Purification using Antibody-Coupled Beads)

1. Buffer Preparation:

   • Filter all water and buffers before use.

   • Equilibration/Wash Buffer: 0.15 M NaCl, 20 mM Na₂HPO₄, pH 7.0

   • Elution Buffer: 0.1 M Glycine, pH 3.0

   • Neutralization Buffer: 1 M Tris-HCl, pH 8.5

2. Bead Pre-equilibration:

   • Resuspend coupled beads thoroughly. Transfer the calculated volume (e.g., 100 μL) to a microcentrifuge tube.

   • Place on magnetic rack for ~1 min until solution clears. Aspirate supernatant.

   • Remove tube from rack. Add an equal volume (e.g., 100 μL) of Equilibration Buffer. Pipette mix 5 times.

   • Place on magnetic rack for ~1 min. Aspirate supernatant.

   • Repeat equilibration wash twice (3 washes total).

3. Sample Binding:

   • Add sample solution to the pre-equilibrated beads. Vortex-mix briefly.

   • Incubate with end-over-end rotation (or gentle manual tube inversion) at room temperature for ≥30 min (adjust based on binding efficiency).

   • Place on magnetic rack for ~1 min. Aspirate and discard supernatant.

4. Washing:

   • Add 5x initial bead volume (e.g., 500 μL) of Wash Buffer. Vortex-mix to resuspend.

   • Incubate with mixing for 1-2 min.

   • Place on magnetic rack for ~1 min. Aspirate and discard supernatant.

   • Repeat wash twice (3 washes total).

5. Elution:

   • Add 3-5x initial bead volume (e.g., 300-500 μL) of Elution Buffer. Pipette mix 5 times.

   • Incubate with end-over-end rotation (or gentle manual tube inversion) at room temperature for 5-10 min.

   • Place on magnetic rack for ~1 min. Transfer the clear supernatant (elution fraction) to a clean tube. This contains the target protein.

6. Neutralization:

   • Add 1/10th volume of the elution fraction of Neutralization Buffer to the elution fraction. Mix immediately to adjust pH to 7.0-8.0.

7. Bead Regeneration & Storage:

   • Add 1 mL Elution Buffer to the beads. Vortex-mix. Place on magnetic rack for ~1 min. Aspirate supernatant.

   • Repeat elution wash once (2 washes total).

   • Add 1 mL Equilibration Buffer. Vortex-mix. Place on magnetic rack for ~1 min. Aspirate supernatant.

   • Resuspend beads in Storage Solution (Section I) at 10 mg/mL. Store at 2-8°C.

Specifications

Especificaciones y pureza

BioReagent, 10 mg/mL

Estabilidad y almacenamiento

Store at -20℃ long term (24 months).

Condiciones de almacenamiento de almacenamiento

Store at -20°C

Enviado en

Ice chest + Ice pads

Este producto requiere envío en cadena de frío. Los servicios terrestres y otros servicios económicos no están disponibles.

Grado

BioReagent