PureCube Epoxy Activated MagBeads available as off-the-shelf items. Our careful in-house production provides low lot-to-lot variation, making activated MagBeads also suitable for commercial production. PureCube Epoxy Activated MagBeads carry an epoxy functionality. Proteins can bind covalently via free amines or thiol groups. The beads are provided as a 25% suspension.
PureCube Epoxy Activated MagBeads provide:
||Coupling of biomolecules via free amine or thiol groups
||Covalent coupling to free amine and thiol groups
||Delivered as a 25 % suspension
|Epoxy Group density
|| higher than 20 µmol/ml
||Activated Epoxy groups
- Sodium dihydrogen phosphate
- Sodium chloride
- Sodium acetate trihydrate
- Sodium hydroxide (NaOH)
- Acetic acid
- Optional: Sodium carbonate monohydrate
- Optional: Ethanolamine
- Magnetic holder for microcentrifuge tubes (for separation of magnetic beads)
- Microcentrifuge tubes (2 mL)
- Centrifuge for 15 mL tubes
- Centrifuge tubes (15 mL)
- End-over-end mixer or thermomixer
|A.||Epoxy coupling protocol:|
- Transfer 1 mL PureCube Epoxy-Activated MagBeads (25%) into a 2mL microcentrifuge tube. Tip: The coupling reaction can be linearly scaled up and down, by increasing or decreasing the amounts of buffers and solutions described in this protocol.
- Place the tube on a magnetic stand and allow the beads to separate. Remove the supernatant.
- Wash the Activated MagBeads four times with 1 ml double distilled water, then wash once with 1 ml coupling buffer I or II.
- Resuspend the MagBeads in 500 µl coupling buffer and incubate for 10 minutes at 20 or 37°C, depending on the stability of your protein. Tip: When coupling a particular protein for the first time, try 3-5 different protein concentrations to make sure you are offering enough protein in the reaction but not wasting any protein.
- Add the protein to be coupled to the mix. The exact protein amount needs to be optimized, and 5 to 10 mg protein is a good starting point. Tip: Choose the highest temperature the protein is stable at. Coupling efficiency increases with temperature, incubation time, and protein concentration.
- Incubate at 20-37°C for 24 hours on an end-over-end shaker or thermoshaker.
- Remove the supernatant and analyze the supernatant in a spectrophotometer. Record absorption at 280 nm to monitor coupling efficiency. Tip: Monitoring absorbance at 280 nm tells you about the coupling efficiency of the protein (compare A280 of the original protein solution to the supernatant in step 6 to determine % coupling). It also helps you identify the optimal amount of protein required for efficient coupling.
- Wash the magbeads five times with 1 ml double distilled water and once with 1 ml Storage buffer. Store at 4°C.
- Optional: Add 500 µl Quenching Buffer and incubate again for 8 h at 20-37°C. Wash five times with 1 mL double distilled water each and once with 1 ml Storage buffer. Note: The quenching step ensures that no free Epoxy groups are left on the MagBeads that might interfere with subsequent assays.
- . Resuspend the coupled MagBeads in 1 mL Storage buffer, yielding a 50% suspension. Store at 4°C.