Filtration in BioProcess – Tangential Flow Filtration – TFF

Filtration is the process of separating the components of a solution based on their size and charge differences using a filter. It is a pressure driven process. Filtration is an important process in biopharmaceutical industry which is used right from the separation of cell mass to the final concentration of the product.

Types of Filtration
Based on the fluid flow, filtration can be of two different types:

(1) Dead End filtration
(2) Cross flow filtration

In Dead end filtration, the fluid flows directly towards the filter under the influence of pressure. While using the dead-end filtration method, all the smaller particles passes through the membrane and all particles those are larger than the pore size of the filter accumulates at its surface. The size of those particles prevents them from passing through the filter medium. These trapped particles start to build up a “filter cake” on the surface of the filter, which reduces the efficiency of the filtration process.

The process is often called as Normal Flow Filtration (NFF). The terms “Normal” and “Dead End” indicate that the fluid flow is perpendicular (normal) to the filter surface towards the end and there is no recirculation of the feed.

Tangential Flow Filtration

Cross-flow or tangential flow filtration is based on the pressurized flow of the fluid flowing tangentially over the surface of the filter membrane, with a portion of the feed pushed through the filter and the remaining being swept sweeping tangentially along the membrane to exit the system without being filtered. The pressure applied forces a portion of the fluid through the filter to the filtrate side.
TFF generally involves the circulation of the fluid parallely over the surface of a filter under pressure. i.e., fluid is swept over the surface of the filter as filtrate is pushed to pass through the filter membrane by the influence of the pressure applied on the feed. The particles which are of greater size than the pore size are swept along with the tangential flow of the fluid. Thus the tangential flow of the fluid prevents accumulation of particles on the flow path of the filtrate. This process has got its name “tangential” by the direction of the fluid relative to the filter.

The portion of the feed fluid which passes through the filter is referred to as filtrate or permeate and that which does not pass through the filter is referred to as retentate. The retentate is again sent to the reservoir to be recirculated.
In TFF equipment, a pump is used to generate pressure on the feed flow. During each pass of the feed fluid over the surface of the filter, the applied pressure forces a portion of the fluid through the filter to the filtrate stream.

Tangential Flow Filtration System Schematic

Applications of TFF
For large-scale industrial filtration, cross-flow or tangential-flow filtration is commonly employed. TFF is mainly used for concentration, diafiltration (buffer exchange, desalting) and separation of biomolecules of varying size measuring from hundreds to thousands of Daltons. TFF can also be used for the clarification or removal of biomass as well as cell debris measuring microns in size from fermentation broth and cell culture broths.

Concentration
It is a simple process which involves the removal of a portion of fluid containing molecules which are smaller than the membrane pore size while retaining the bigger molecules in retentate. The concentration of the fluid increases proportionately as the filtrate is removed. If the volume decreases by a factor of 2, the concentration of the fluid doubles.

Diafiltration (DF)
It is the separation process that washes smaller molecules through the filter leaving behind the larger molecules in the retentate without changing the concentration. It can be used to remove spent media in fermentation/cell culture broths, remove salts or exchange buffers in protein suspensions. This process can be used to enhance either purity or yield.

UltraFiltration schematic

In Diafiltration process, buffer is added into the feed tank while filtrate is removed continuously from the process. In processes where the products of interest are in the retentate, diafiltration washes and removes other components out of the retentate into the filtrate, thereby exchanging buffers and reducing the concentration of unwanted materials. In cases where the products are in the filtrate side, DF process washes them through the filter membrane into a collection tank.

Diafiltration can be done in the following two different modes

Batch Diafiltration
In Batch DF, the process fluid is first diluted with a large volume of diafiltration buffer and then the retentate is concentrated to the original volume or upto certain concentration of biomolecules in the retentate. Once this concentration is reached, another volume of buffer is added. This cycle is repeated until required concentration of biomolecules in the retentate is reached or until the desired total volume buffer is added.
In this method, buffer exchange is not efficient as the retentate concentration decreases as the buffer is added and increases during concentration.


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Continuous Diafiltration
In Continuous DF, the diafiltration buffer is added to the feed tank at the same flow rate at which the filtrate is removed. By this method, the volume of the fluid in the process can be kept constant while the smaller molecules which can permeate through the filter are washed away in the filtrate. This requires some level control for DF buffer addition to keep the retentate level constant.
Continuous DF requires less buffer volume and less filtrate removal than Batch DF to achieve the same extent of washing.

Process parameters in TFF
Transmembrane Pressure (TMP) – The pressure acting across the membrane which drives the fluid to the filtrate side carrying all the permeable molecules.
Transmembrane Pressure (TMP)

Pin – Inlet Pressure i.e., on the Feed side
Pout – Outlet Pressure on the retentate side
Pp – Pressure in the permeate line

Cross flow velocity The flow rate of the fluid through the feed line and across the filter membrane surface. This is the force required to sweep away the molecules which tend to accumulate on the filter and cause fouling thereby restricting the filtrate flow.

Pressure Drop (ΔP) The pressure difference along the narrow feed channel across the membrane from the feed to the retentate.

Pressure Drop (ΔP).

Membrane Area The effective filtration area of the membrane required for filtration of a sample at a specified time can be calculated as
Membrane Area

V – Total Volume of filtrate collected (liters)
T – Process Time (hour)

Permeate flux (Fp) – The flow rate of the permeate/filtrate per area of the membrane through which it is passing. It is measured as liters per m2 per hour or LMH.
Permeate flux

Qp – Permeate flow rate (L/H)
A – Area of the membrane (m2)

Normalized Water Permeability (NWP) The rate of clean water flux through the membrane under conditions of standard pressure and temperature. NWP is used for measuring the effect of cleaning of filter membranes. The water flux is measured as liters per m2 per hour and NWP is expressed as LMH/bar at 250C.
Normalized Water Permeability (NWP)

K – Temperature Correction Factor
K = µT Deg C/µ25 Deg C
where, µT Deg C is the Viscosity of water at a given temperature and µ250C is the Viscosity of water at 250C.

Types of TFF
TFF can be divided into categories based on the size of particles being filtered.

Micro filtration (MF)
It is a low-pressure process for separating fluids having particles of 0.05-1micron in size. MF is mostly used in the upstream of a process where cells or cell lysate has to be separated from the fluid. It is mainly used for clarification of biomass from fermentation/cell culture broths.

Ultra filtration (UF)
It is a selective separation process which utilizes a high pressure of up to 10 bar. UF is used for concentration, desalting, buffer exchange or separation of proteins from buffer components based on the molecular weight of the proteins. Depending on the molecular weight of the proteins to be separated, membrane pore size may vary from
1 kD to 1000 kD.

Nanofiltration (NF)
NF is a special process which uses very tight membranes to remove salts, minerals and other smaller molecules having low molecular weights from water and other solvents. The membrane pore size may be as low as 1 kD or less than that.

Virus filtration
This is a type of UF used for the removal of viruses from the protein molecules or other media components which can be used for virus reduction or virus harvest.

(Sincere thanks to guest editor St. Mane for his contribution)

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