Magnetic beads Dynabeads

If further purification is required add antibody-coated (anti-CD2,-CD14, and -CD19) magnetic beads (Dynabeads M-450) at a ratio of 10 beads/cell. Incubate at 4°C for 30 min with gentle rocking. 

Streptavidin-coated magnetic beads (Dynabeads M280-SA, Invitrogen, 6.7 x 108beads/mL, 1 mg beads binds 700 pmoles free biotin). 

Dynabeads M-270 streptaviclin-coated magnetic beads from Dynal (Oslo, Norway)... 

Tosylactivated magnetic beads (MB-Tosyl) (Dynabeads M-280 Prod. No. 142.03, Dynal Biotech ASA, Oslo, Norway). Hydroquinone (Sigma), hydrogen peroxide (Merck, Germany). Bradford solution (Bio-Rad protein assay Catalog No. 500-0006) was purchased from Bio-Rad laboratories GMGH (Munich, Germany). 

Dynabeads anti E. coli 0157 (Dynal Ltd) 0157 — Magnetic beads AFNOR DYN 16/2-0696. ISO EN 16654... 

The international reference method to detect and recover E. coli 0157 (ISO EN 16654, Fig. 1) inclndes a step of Immuno-Magnetic Separation (IMS), which requires the nse of magnetic beads coated with polyclonal antibodies against 0157 antigen (Dynabeads anti E. coli 0157, Dynal Ltd, Norway). The cell bead complexes are recovered from the medinm by the application of a magnetic field causing the complexes to be concentrated in the tnbe, and enabling the bulk medium to be discarded. The concentrated cells are next plated onto CT-SMAC for strain recovery and confirmation. Chapman et al. (1994) fonnd that IMS was 100-fold more sensitive for detection of E. coli 0157 H7 than direct culture on isolation media. 

Reagents for Magnetic Bead Preparation (Dynabeads Are Typically Used)... 

In a microtube, add 1 mg of M-270 streptavidin-coated Dynabeads to 1 mL TBS and place the tube on a magnet to discard the supernatant. Off the magnet, resuspend the beads in 1 mL of MTBS for blocking nonspecific sites. Place the microtube on a rotating wheel for 1 h at room temperature. The rotation is necessary for keeping the beads in suspension and should be slow. 

Before the antibody incubation is complete, prepare the Dynabeads by gently shaking the container to resuspend the beads. Pipette 75 pi of the Dynabeads solution into a new, sterile, labeled 1.5-ml microcentrifuge tube. Place tube into the magnetic separation stand for 1 min and then pipette off and discard supernatant. Equilibrate the beads by resuspending them with 200 pi lx dilution... 

Selection is carried out by adding the antigenbinding complexes are removed by washing the beads. A magnetic particle concentrator (Dynal MFC) is used to wash and recover the beads carrying bound ARM complexes. 

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