Tissue Homogenizer - Principle, Parts, Types, Procedure

Tissue Homogenizer is a laboratory device that is used to break biological tissues into small and uniform particles.

It physically disrupts the tissue structure by rupturing the cell wall and cell membrane. During this process, intracellular components like proteins, nucleic acids (DNA and RNA) and microorganisms are released from the tissue matrix.

The uniform mixture formed after this process is called homogenate. It is used for proper sample preparation in cancer research, neuroscience, molecular biology and other life science studies. Tissue homogenizer may work by mechanical shearing, pressure, ultrasonic sound waves or bead milling.

Working Principle of Tissue Homogenizer

Working Principle of Tissue Homogenizer is based on the physical disruption of tissue sample by applying mechanical or acoustic force.

The tissue is broken into small particles and uniform mixture is formed. During this process, cell membrane and cell wall are ruptured. The intracellular components such as proteins, DNA, RNA and microorganisms are released from the tissue matrix.

In rotor-stator homogenizer, the sample is pulled between rapidly rotating rotor and stationary stator. Strong shear force is produced in the narrow space. This force breaks the tissue and forms fine suspension.

In bead mill homogenizer, the tissue sample is placed with small beads in a closed tube. The tube is shaken rapidly. The beads strike the tissue and break the cells by collision and impact.

In ultrasonic homogenizer, high frequency sound waves are used. These waves produce cavitation. In this process, small bubbles are formed and collapsed, which produce shock waves and rupture the tissue.

In high pressure homogenizer, the sample is passed through a very small gap under high pressure. Shear force, turbulence and cavitation are produced. These forces reduce the size of tissue particles.

In manual tissue grinder, the tissue is crushed between pestle and glass tube. It breaks the sample by crushing and shearing.

Thus, the tissue homogenizer works by breaking the tissue structure and releasing the cell contents. The uniform mixture formed is called homogenate.

Parts of a Tissue Homogenizer

The following are the important parts of Tissue Homogenizer–

Motor or Drive Unit– It is the powered part which rotates the shaft at high speed.
Rotor– It is the inner metal shaft that rotates very fast and produces suction and shear force.

Stator– It is the stationary outer case present around the rotor, which contains narrow slots or openings.
Generator Probe– It is the assembly containing both rotor and stator, and it is placed inside the tissue sample.
Tubes or Vials– These are sealed containers that hold the tissue sample and grinding beads.

Beads or Grinding Media– These are small round particles which collide with tissue and break the cells. They may be made up of glass, ceramic, stainless steel, zirconium, silica or garnet.
Agitator Mechanism– It is the mechanical part that shakes or vibrates the tubes at high speed.
Generator– It converts electrical energy into high frequency mechanical vibration in ultrasonic homogenizer.

Probe or Horn– It is a vibrating rod, mostly made of titanium, which is dipped into the liquid sample and produces cavitation.
High-Pressure Piston Pump– It draws the sample and pressurizes it with the help of electric motor.
Homogenization Device or Valve– It is the part where actual homogenization takes place through a very small gap.

Dampers– These are placed on suction and outlet pipes to reduce pulsation, vibration and noise.
Mortar or Tube– It is the outer glass container that holds the tissue sample in manual homogenizer.
Pestle– It is a blunt rod like part which crushes and grinds the tissue against the wall of the tube.

Operating Procedure of Tissue Homogenizer

The following are the operating procedure of Tissue Homogenizer–

The tissue sample is kept on ice and solid tissue is cut into very small pieces of about 1 mm size by sterile scalpel or razor.
The cut tissue is placed in a tube and chilled sterile buffer is added into it.
Protease inhibitors and phosphatase inhibitors are added to prevent the breakdown of proteins during homogenization.

In probe based homogenizer, the probe is completely dipped in the liquid sample to avoid foaming and air mixing.
The homogenizer is operated for short bursts of about 10 to 15 seconds and the sample is kept on ice during this process.
Heat is produced during homogenization, so the tube is kept in wet ice to protect the sample components.

After tissue disruption, the homogenate is centrifuged at 14,000 to 16,000 × g for 10 minutes at 4°C.
The solid cellular debris settles at the bottom and forms a pellet.
The liquid supernatant is carefully collected, which contains the target intracellular components.

The probe is immediately cleaned with water and suitable solvent like ethanol or methanol to prevent contamination of next sample.

Types of Tissue Homogenizer

The following are the different types of Tissue Homogenizer–

Bead Mill Homogenizer– It uses rapidly moving small beads such as glass, ceramic or metal beads inside a sealed tube. The tissue is broken by impact and friction of beads.
Rotor-Stator Homogenizer– It has a fast rotating rotor inside a stationary stator. The sample is pulled inside and tissue is disrupted by suction and high shear force.

Ultrasonic Homogenizer– It is also called sonicator. It uses high frequency sound waves through a vibrating probe. These waves produce cavitation and cells are ruptured by shock waves.
High-Pressure Homogenizer– It forces the sample through a very small opening or valve under high pressure. The particles are reduced by turbulence, cavitation and pressure drop.
Microfluidizer Homogenizer– It is a special type of high pressure homogenizer. The sample is passed through fixed geometry interaction chamber for uniform particle size reduction and dispersion.

Manual Homogenizer– It includes mortar and pestle, Dounce homogenizer and Potter-Elvehjem grinder. The tissue is crushed and sheared manually between a container and pestle.
Paddle Blender– It is also called stomacher. It uses mechanical paddles to squeeze the sample inside a bag against a solid surface. It releases cells or microorganisms into solution without complete blending of tissue.

Applications of Tissue Homogenizer

The following are the applications of Tissue Homogenizer–

It is used for disruption and lysis of cells to release intracellular components such as DNA, RNA, proteins and organelles.

It is used for recovery of microorganisms from tissue sample and tissue matrix.
It is used to expose antigens for immunological assay and immunohistochemistry.
It is used to grind and prepare different samples like animal tissue, plant material, bone and soil.

It is used in pharmacology, forensics and environmental testing for sample preparation.
It is used in pharmaceutical work for preparation of stable drug suspension and vaccine adjuvants.
It is used in the preparation of lipid nanoparticles and targeted drug delivery system.

It is used in food and beverage industries for making stable and uniform emulsions like milk, juices and sauces.
It is used in cosmetic production for mixing and preparation of uniform creams, gels and lotions.
It is used in material science and nanotechnology for particle size reduction and deagglomeration.

It is used for formation of nanoemulsions and nanoparticles.
It is used in special laboratory works like DNA shearing, chromatin shearing and homogenization of samples under cryogenic condition.

Advantages of Tissue Homogenizer

The following are the advantages of Tissue Homogenizer–

It converts heterogeneous tissue sample into uniform and stable mixture.
It gives consistent sample preparation and reduces variation in experimental result.

It ruptures the cell wall and cell membrane and releases intracellular components like proteins, DNA, RNA and microorganisms.
It makes the intracellular components available for further analysis.
It saves time and manual labour than traditional hand grinding method.
It can process many samples at a time in some types like bead mill homogenizer.

It reduces the chance of contamination because many homogenizers use sealed and single-use tubes.
It also reduces aerosol formation of infectious agents during sample processing.
It can be used for many types of samples like soft tissue, liquid sample, bone, skin and seeds.

It can work by different mechanisms like mechanical shear, ultrasonication and bead milling.
It increases the recovery of microorganisms and target analytes from tissue matrix.
It improves the sensitivity of diagnostic tests by increasing the amount of target materials.

In food, cosmetics and pharmaceutical industries, it improves emulsion stability and prevents precipitation.
It improves texture, appearance and flavour of industrial products.

Limitations of Tissue Homogenizer

The following are the limitations of Tissue Homogenizer–

It produces heat during homogenization, mainly in rotor-stator homogenizer, ultrasonic homogenizer and high-pressure homogenizer.
The heat produced during process may damage heat sensitive proteins and enzymes.

Some homogenizers can process only small amount of sample, usually few milliliters or grams.
Bead mill homogenizer, manual tissue grinder and French press are not easily used for large scale work.
There is chance of sample contamination by grinding materials, such as bead particles in bead mill and titanium particles from sonicator probe.

Probe based homogenizer may cause cross contamination between samples when it is not cleaned properly.
Some homogenizers produce high noise during operation, mainly ultrasonic homogenizer and bead mill homogenizer.
Foaming and air mixing may occur in blender and rotor-stator homogenizer due to vortex formation.

Air bubbles may cause protein denaturation and uneven lysate formation.
High pressure homogenizer and microfluidizer are costly and large in size.
Moving parts like homogenizing valve, seals and ultrasonic horn are worn during use and need regular maintenance.

Manual homogenizer and probe based rotor-stator homogenizer usually process one sample at a time.
Ultrasonic homogenizer is not suitable for solid tissue and highly viscous liquid sample.
In viscous sample, sound waves cannot pass properly and cavitation becomes difficult.

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Tissue Homogenizer – Principle, Parts, Types, Procedure