A centrifuge is an equipment that applies the principle of centrifugation to separate particles from the mixture. The particles in the mixture separates based on their density, i.e., the denser particle settles down as pellets in the container, and the lighter particle remains suspended as supernatant in the mixture.
Centrifugation separates particles based on their size, shape, density, and viscosity by centrifugal force. Centrifugal force is the force generated by the rotation of motors. The following factors influence the centrifugation:
- The density of the samples: The denser particles settle down, and particles with less density float or remain suspended in the solution.
- The viscosity and temperature: The spinning of motors causes an increase in temperature, and increased temperature can disrupt the bonds and stability of the particles helping in the separation of the particles. Similarly, the larger the sample’s viscosity, the easier it is to separate its component.
- The distance of particle displacement: The greater the distance of particle displacement, the greater the separation rate.
- The speed of rotation: The higher the speed, the faster the particles separate.
The acceleration applied to the sample is called relative centrifugal force (RCF) or G-force. The particles will move away from the axis of rotation when the RCF exceeds the frictional force.
Parts of a Centrifuge
The centrifuge machine has the following parts:
It is in the center that is very powerful and creates a spin.
The rotor is attached to the motor. The rotation occurs at no angles (horizontal centrifuge) or different angles; 45° (fixed angle centrifuge) and 90° (vertical centrifuge). Depending on the centrifuge, the loading of the sample may be done at a fixed angle, i.e., angle of rotation, or the container will adjust itself to a different angle during operation. The second method is also known as swinging bucket and is commonly found in horizontal centrifuges.
Swinging bucket rotors: These are the rotors present in centrifuge that allows the container to change the angle while in motion. The samples move through the density gradient without interruption due to the parallel geometry of the bucket.
These containers hold tubes with the materials/sample and it rests on the rotor.
The types of control vary based on the centrifuge selected. Some are preprogrammed, and others are entirely customized with digital display. Regardless of the type of control, the centrifuge will run the motor based on the provided settings.
Operation of Centrifuge
The operation of the centrifuge is easy. Steps of operating centrifuge:
- Place the test tube with the sample into the container/portals.
- Balance the samples, but insert water-filled tubes for balance when there is an odd number of samples.
- Close the lid and select the required time and speed.
- Start the centrifuge and wait for the cycle to complete.
- When the centrifuge stops, take out the balance and the sample. The separated sample is now ready for analysis.
Why is balancing necessary?
The centrifuge runs by force from the rotor’s spinning, creating centrifugal force or G-force. The force magnifies the slight weight difference between samples but magnifies any differences if the weight distribution is uneven. Hence, if there is a little weight difference between samples, the machine will vibrate/shake uncontrollably. So, balancing the centrifuge is very necessary.
How to balance a centrifuge?
The ways to balance the centrifuge are as follows:
- Ensure that the tubes are evenly filled with liquid with a similar density.
- Make sure that the masses in the tubes are within 0.1 grams of each other.
- Place the tube just opposite one another to keep the gravity in the center.
- Use water to balance when the samples are in odd numbers.
Types of Centrifuge
There are different types of centrifuges based on optical detection, size, refrigeration capability, and rotor speed. The details of each type should be studied while choosing the perfect centrifuge.
Benchtop centrifuges are popular for their characteristic feature of requiring small space on the benchtop. Other features of benchtop centrifuges are the speed (RCF) of this centrifuge; it ranges from a few hundred to 50,000 x g and has interchangeable rotors; fixed angle, continuous flow, and swinging bucket rotors are available. Tubes of benchtop centrifuges range from less than 1 ml to a few liters.
As the name suggests, the microcentrifuges accommodate tubes containing small/micro volumes of samples like 2 ml, 1.2 ml, 0.5 ml, and PCR (polymerase chain reaction) tubes. So, these apply most frequently in the microbiological laboratory for separating nucleic acid and proteins. It typically spins at speeds up to 16,000x g, but in the case of specialized forms, the rate can reach up to 30,000 x g. Some models also have interchangeable rotors and tube adaptors.
Vacuum centrifuge uses centrifugal force, vacuum, temperature, and gas to remove liquid or gas from the sample to concentrate or desiccate the samples. Purifying nucleic acids, proteins, peptides, and other components used in research laboratories are some its uses.
The refrigerated centrifuge works at a constant temperature with maximum speed. The temperature range of the centrifuge is between -20 to -30℃. Centrifugation of samples like DNA, RNA, and antibodies, which need low but constant temperature, is easy in this machine.
These are also used for samples that need to be stored at a consistent temperature. These centrifuges must function at maximum speeds while maintaining a steady temperature.
The acceleration speed of ultracentrifuges is typically up to 1,00,000 x g, but the rate can reach 2,00,000 x g.
Analytical centrifuges have different features that allow samples to detect as they spin in real-time. A light-based optical system like the light adsorption system, Rayleigh system, and the alternative Schlieren system helps observe the separated materials and monitor the speed.
Handling of Centrifuge
For proper functioning of the centrifuge, it requires constant care and maintenance. There are some measures to consider before, during, and after using the device to increase its lifespan. They are:
Lubricate the machine
The rings in the centrifuge are crucial as they prevent leaking from the sample. Hence, lubricating of rings after cleaning, rotor installation, and repairing with Hettich grease is necessary.
The centrifuges should be checked regularly for any damage, unfamiliar noises, grinding and unnecessary vibration. The damages and dysfunctionality should be repaired regularly.
The rotors, chamber of rotors, the interior of the centrifuge, and electrical components are some of the parts that need constant cleaning using alcohol-based disinfectant and a soft cloth.
It should be ensured that the personnel using the centrifuge know how to operate it well. The tubes are balanced, and adjusting speed and compartment mass are things the operator should know before using the machine.
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