Magnetic beads preparation

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

B. Polymeric Urea [Benzene, diethenyl-, polymer with ethenylbenzene, [[[[(1 methylethyl)amino]carbonyt]amino]methyl] deriv.] A 10.0-g. portion of benzylamine polymer beads prepared as in Part A and 125 ml. of tetrahydrofuran (Note 6) are combined in a 300-ml., three-necked, round-bottomed flask equipped with a magnetic stirrer, a dropping funnel, and a condenser fitted with a gas-inlet tube A nitrogen atmosphere is established in the system, and the slurry is stirred while 1.35 g. (0.0159 mole) of isopropyl isocyanate [Propane, 2-isocyanato-] is added. This causes an exothermic reaction, which subsides after about 20 minutes. The mixture is then stirred at room temperature for 22 hours and at reflux for an additional 4 hours. The beads are collected by filtration, washed with 150-ml. portions of tetrahydrofuran (Note 6) and methanol, and dried under reduced pressure over calcium chloride to yield 9.09 g, of the isopropyl urea polymer. 

Micro- and nanobeads with magnetic properties have recently become popular since these tools can be manipulated, e.g., collected in the region of interest. Magnetite nanoparticles are introduced in order to render the polymeric beads magnetic. Preparation and application of magnetic beads will be discussed in more detail in Sect 5.5. 

Precipitation was found to be a very useful method for preparation of nanobeads with magnetic properties [17] since not only indicators but also small lipophilic magnetite nanobeads (having diameter of a few nanometers) can be incorporated inside the polymeric beads. Such multifunctional magnetic beads can be guided to the region of interest, be collected and manipulated there. 

Beads of cross-linked polystyrene containing magnetite crystals (Fe304, a ferromagnetic material) have been prepared and successfully used for solid-phase synthesis [125-127]. The magnetic beads could be readily separated from the reaction mixture with a magnet. 

Prepare three different pools of modified MB-Tosyl magnetic beads (i) with the anti-sulfonamide class-specific antibodies (ii) with the non-specific antibody (with the non-immunized serum) and, (iii) without any antibody, only treated with the corresponding coupling buffer [2],... 

The magnetic beads with attached oligo(dT) probes can be used for purifying mRNA either from a total RNA preparation or directly from the tissue [30-32], The advantage of the latter is that mRNA can be isolated from very small amounts of tissue or cells. The tissue homogenate obtained... 

Fig. 1. Preparation of ssDNA from biotinylated PCR-amplified DNA. (a) DNA is amplified using one primer that has a biotin molecule (B) at the 5 end and one primer that lacks biotin, (b) Double-stranded amplified DNA is captured on a streptavidin-coated magnetic bead via the biotin molecule, (c) The dsDNA is denatured, with the biotinylated strand remaining attached to the streptavidin-coated magnetic bead and the nonbiotinylated strand being released into the supernatant, (d) The nonbiotinylated strand is used as an ssDNA sequencing template.

Notes. The different brands of streptavidin magnetic beads have different molarities of attached streptavidin, different resuspension protocols, and different capture properties. Bead volumes given in this protocol are for Promega (Madison, WI) paramagnetic particles (No. Z5241) prepared as indicated below. If other beads are used, appropriate volumes may differ by a factor of 3 or more from those presented. 

NH4 ZSM-5 was prepared from NaZSM-5 by ion exchange technique. The sample taken in a conical flask is stirred in NH4NO3 (0.5 M) in solution at 343°K for 7 hrs. with the help of magnetic bead. The zeolite is then filtered, and washed with deionised water. The same procedure is repeated three times. After the final exehange, the sample is washed free of salts and dried in an oven at 373 K for 12 hrs. NH4 ZSM-5 thus obtained is converted to hydrogen form by calcining at 823 K for 7 hrs. using a furnace with a flow of dry air. 

HCCA (0.67 mg/mL in ethanol acetone, 2 1). Dissolve approximately 1 mg HCCA in 500 pL acetone to make a 2-mg/mL stock solution. The matrix usually takes 5-10 min to completely dissolve at room temperature. We usually start the magnetic beads fractionation while waiting for the matrix to dissolve. Then add one part of above stock solution to two parts ethanol (e.g. 100 pL stock solution to 200 pL ethanol). Prepare the matrix solution daily. 

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