Rotor swinging

Basically, rotors come in four varieties fixed-angle rotors, vertical tube rotors, swinging-bucket rotors, and zonal rotors. The first three rotors are discussed below. Zonal rotors will be discussed In a later section. 

Fig. 11. (a) Diffeiential centrifugation (pelleting), where time 1 < time 2 < time 3 < time, (b) Rate 2onal separation in a swinging-bucket rotor, where tube 1 represents the density gradient solution, tube 2 the sample plus the gradient, and tube 3 the separation of sample particles under a centrifugal force, where... 

Pin MiUs In contrast to peripheral hammers of the rigid or swing types, there is a class of high-speed mills having pin breakers in the grinding circuit. These may oe on a rotor with stator pins between circular rows of pins on the rotor disk, or they may be on rotors operating in opposite directions, thereby securing an increased differential of speed. See also the Mikro-ACM pulverizer described later. 

Add 30 mL of 15-50% (w/w) gradient solution to the ultracentrifuge tubes of the swinging bucket rotor (max vol 40 mL). Sucrose concentrations increase from the top to the bottom of the tube. 

Ultracentrifuge the tubes at 84,000 for 17 h in a swing rotor (Hitachi, RPS27-2, Tokyo, Japan). Note that at the beginning and end of centrifugation, acceleration and deceleration rates should be low so as not to disturb the gradient of the tubes. 

Step 1. Load a disposable mini-chromatography col-umn with Sephadex G-50 (fine) dextran that was previously permitted to equilibrate with 5 volumes of the desired new buffer. Allow all the liquid to drain under the earth s gravitation to leave a 1 mL bed volume. Transfer this spin-column to an appropriately sized centrifuge tube and spin at 100 X g (calculated for the radial distance to the tip of the column itself, and not the bottom of the centrifuge tube) for 2 min in a swinging bucket rotor". The volume of the now semi-dry gel bed will be reduced by 25-40%, and the media will appear as a gel mass that may even break away from the walls of the chromatography tube. (This is normal and will not affect performance.)... 

The tubes of the swinging bucket rotor type are accommodated in a pivoted bucket moving from vertical to horizontal position during acceleration under the influence of the centrifugal field. The sedimentation path length is lowest in vertical rotors and highest in swinging-bucket rotors. 

Fig. 5.2. rpm-g graph for fixed-angle (type 45 Ti) and swinging-bucket (SW 28) Beckman Coulter ultracentrifuge rotor... 

To extract the liberated fatty acids, 1.5 ml of 0.1 mM carbonate-bicarbonate buffer, pH 10.5 is added and the mixture is shaken for 10 s. Centrifugation for 45 min at 1500 xgin a swing-out rotor will separate the water from the lower organic phase. In a scintillation vial, 2 ml of the upper water phase are mixed with 50 pi of glacial acetic acid containing 500 pg ferric acid before the scintillation liquid is added (16 ml of Ecoscint toluene (7 1 volume). Liquid scintillation counting is done for 5 min and the LPL activity is calculated from the difference in counts between the blank and the sample vials [40]. 

Most laboratories have a standard low-speed centrifuge used for routine sedimentation of relatively heavy particles. The common centrifuge has a maximum speed in the range of 4000 to 5000 rpm, with RCF values up to 3000 X g. These instruments usually operate at room temperature with no means of temperature control of the samples. Two types of rotors, fixed angle and swinging bucket, may be used in the instrument. Centrifuge tubes or bottles that contain 12 or 50 mL of sample are commonly used. Low-speed... 

Rotors for a high-speed centrifuge. A Fixed angle B swinging bucket C vertical. Courtesy of Beckman Instruments, Inc. 

Operation of a swinging-bucket rotor. A Loading of sample. B Sample at start of centrifugation. C Sample during centrifugation separates into two components. D Rotor at rest. 

---