The DH92-IIN ultrasonic cell disruptor is a multifunctional and multi-purpose instrument that uses strong ultrasound to generate cavitation effect in liquid and treat substances with ultrasound. It can be used for the fragmentation of various animal and plant cells, virus cells, as well as for emulsification, separation, homogenization, extraction, defoaming, cleaning, and accelerating chemical reactions. Widely used in fields such as biochemistry, microbiology, medicinal chemistry, surface chemistry, physics, zoology, etc.

◆ Used for crushing animal and plant cells, bacteria, spores or tissues, extracting proteins from cells, and conducting scientific cultivation experiments for viruses and vaccines.

Accelerate the reaction rate of chemistry, physics, etc. and accelerate the degassing of liquids.

Research analysis on crude oil dilution, oil-water emulsification, accelerated de crystallization, and glass homogenization.

◆ Disperse rare earths, various inorganic minerals, and prepare a homogenized mixture of latex particles with a concentration of nearly 1/100 nanometer.

Quick and powerful high-precision cleaning solution for micro holes and blind holes in molds.

Experimental purpose

1. Understand the basic principles of ultrasonic cell lysis

2. Master the basic techniques of ultrasonic cell lysis

Experimental principle

When strong sound waves act on a solution, the cavitation phenomenon of bubble generation, growth, and fragmentation is related. The shock wave and shear force caused by cavitation cause cell lysis.

Materials and reagents

Bacteria 10ml, beaker, Shaved ice, 75% alcohol cotton ball.

Methods of cell lysis:

1、 Mechanical fragmentation method: refers to the use of crushers, grinders, homogenizers, etc. to break down cells.

1. High speed organization crushing: Mix the material into a thin paste like liquid, place it in a cylinder with about 1/3 volume, cover the cylinder tightly, turn the speed controller to the slowest position first, turn on the switch, and gradually accelerate to the desired speed. This method is applicable to animal visceral tissues, plant fleshy seeds, etc.

2. Glass homogenizer homogenization: First, place the shredded tissue in a tube, then insert it into a grinding rod and grind it back and forth. Move it up and down to crush the cells. This method has a higher degree of cell fragmentation than high-speed tissue crushers and is suitable for small quantities and animal organ tissues.

2、 Physical fragmentation method: refers to the use of temperature, pressure, or ultrasonic waves to break cells apart.

1: Treat cell suspension with ultrasound of a certain power to cause rapid shaking and rupture of cells (breaking cell walls and organelles with the vibration force of ultrasound).

Mechanism: It may be related to the cavitation phenomenon caused by the generation, growth, and fragmentation of bubbles when strong sound waves act on the solution. The shock wave and shear force caused by cavitation cause cell lysis.

The efficiency of ultrasonic fragmentation depends on factors such as sound frequency, acoustic energy, processing time, cell concentration, and cell type. (Pay attention to cooling during use to prevent overheating).

2. High pressure fragmentation: The cell suspension is sprayed from the annular gap of the high-pressure chamber onto the stationary impact ring, and is forced to change direction and flow out through the outlet pipe. During this process, cells undergo high-speed shear collisions and changes from high pressure to normal pressure, thereby breaking down and releasing their contents.

This is an ideal method for gently and thoroughly breaking down cells.

3. Repeated freeze-thaw method: Freeze the cells below -20 degrees Celsius, thaw at room temperature, repeat several times. Due to the formation of ice particles inside the cells and the increase in salt concentration in the remaining cell fluid, swelling occurs, causing the cell structure to break down.

3、 Chemical fragmentation method: refers to the use of organic solvents or surfactants such as formaldehyde and acetone to act on the cell membrane, causing the structure of the cell membrane to be damaged or the permeability to change.

Some animal cells, such as tumor cells, can be disrupted by cell membranes using sodium dodecyl sulfate (SDS), sodium deoxycholate, and other agents. The concentration is generally 1mg/ml.

4、 Enzymatic fragmentation method: Select appropriate enzymes to destroy the cell wall, and then break down the protoplasts in a hypotonic solution.

Bacterial cell walls are thicker and can be treated with lysozyme for better results.

Standard formula for cracking solution: 50mM Tris HCl (pH 8.5-9.0), 2mM EDTA, 100mM NaCl, 0.5% Triton X-100, 1mg/ml lysozyme. Lysozyme works better within this pH range.

Comprehensive description: Regardless of which method is used to crush tissue cells, it will release intracellular proteins or nucleic acid hydrolytic enzymes into the solution, causing the biodegradation of large molecules and reducing the mass of natural substances. Adding diisopropylfluorophosphate (DFP) can inhibit or slow down autolysis; Adding iodoacetic acid can inhibit the activity of proteolytic enzymes that require hydrophobic groups in their active centers. Adding phenylsulfonyl fluoride (PMSF) can also remove the activity of proteolytic enzymes, but not all of them, and should be added several times while crushing; In addition, pH, temperature, or ionic strength can be selected to make these conditions suitable for the extraction of the target substance.

Precautions before use:

1. Select an appropriate probe based on the volume of the sample being processed, and wipe the probe with an alcohol cotton ball before and after use. To replace the probe, use a specialized wrench;

When the AMPLITUDE knob is turned on, the probe should not be exposed to air, and in special circumstances (test probe), it should not exceed 10 seconds;

3. Prepare an ice box before use, and the sample must be placed in an ice bath during processing. The sample concentration should not be too high.