# US 5,704,889 A · Spillover collection of sparse components such as mononuclear cells in a centrifuge apparatus

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- Number: US 5,704,889 A
- Document ID: US5704889A
- Status: Granted
- Kind: A
- Application number: 08/422,598
- Filed: 1995-04-14
- Granted: 1998-01-06
- Earliest claimed: 1995-04-14
- Assignees: Cobe Laboratories, Inc.
- Inventors: Dennis Hlavinka; Thomas J. Felt
- Web page: https://patentgrep.com/patent/US5704889A

## CPC

- B04B 5/0442: {with means for adding or withdrawing liquid substances during the centrifugation, e.g. continuous centrifugation}
- B04B 2005/045: {having annular separation channels}
- B04B: CENTRIFUGES (high-speed drum mills B02C19/11; domestic spin driers D06F; analysing, measuring or monitoring physical or chemical properties of samples during centrifuging, see the relevant subclasses for these procedures, e.g. G01N)

## Abstract

A centrifuge apparatus is used for collecting white blood cells (WBC), primarily mononuclear cells, from whole blood stratified into layers. A thin mononuclear (MNC) layer is formed at the interface of red blood cells and plasma. A barrier is positioned in the separation vessel of the centrifuge at a location to intercept the thin layer. MNC fluid is allowed to pool behind the barrier before collection is started. To collect the MNC pool, the stratified red blood cell layer is raised from below the interface level by slowing or reversing flow in the RBC exit line thereby causing the MNC pool to spill over the barrier into a well in which a collect line is positioned. Collection ceases when a desired percentage of the pool is removed and the normal position of the interface is re-established; thereafter the pool builds again. By raising the MNC pool from below, improvements in purity and collect volume are achieved. The collection procedure can be useful for harvesting granulocytes and, in general, any sparse stratified component of a centrifuged solution where the sparse component is layered between more dense and less dense strata.

## Claims

### Claim 1

The centrifugal method of harvesting a sparse component within a liquid where the sparse component is stratified in a first layer located between a second layer of more dense component and a third layer of less dense component, an interface formed at the junction of said third layer with the layers of more dense components, said method comprising the steps of:
providing a separation vessel for use with centrifuge apparatus, said vessel having a barrier located therein, an inlet line for delivering said liquid to said vessel, a collect line for collecting said sparse component, a first exit line for exiting said more dense component, and a second exit line for exiting said less dense component;
providing a control system so that said apparatus can be operated to collect said sparse component, including an accumulation phase and a spillover phase;
during the accumulation phase, providing for establishing the level of said interface in a steady state condition at said barrier to allow said sparse component to form a pool while said less dense component flows past said barrier to said second exit line and said more dense component exits said vessel through said first exit line;
during the spillover phase, providing for raising the level of said interface from below the level of said interface to cause said pool to spill past said barrier to said collect line for harvesting said sparse component.

### Claim 2 (depends on 1)

The method of claim 1 wherein the step of raising the level of said interface is accomplished by halting the delivery of liquid through said inlet line to reverse the flow of said more dense component in said first exit line and wherein a port for said first exit line is provided in said vessel below the level of said interface.

### Claim 3 (depends on 2)

The method of claim 2 wherein the step of raising the level of said interface includes adding said more dense component to said vessel at a significant distance from said barrier to minimize disturbance of said pool.

### Claim 4 (depends on 3)

The method of claim 3 wherein the step of establishing the level of said interface in a steady state condition is accomplished by monitoring the level of said interface at a significant distance from said barrier to maintain the level of said more dense component at the monitoring location and allowing the formation of a pool of said sparse component at said barrier.

### Claim 5 (depends on 4)

The method of claim 4 wherein said pool is accumulated by locating said inlet line at a significant distance from said barrier wherein said sparse component is separated from said liquid by centrifugal action by the time the liquid flow reaches the location of said barrier.

### Claim 6 (depends on 5)

The method of claim 5 wherein said liquid is whole blood, said sparse component is essentially white blood cells, said less dense component is essentially plasma, and said more dense component is essentially red blood cells.

### Claim 7 (depends on 6)

The method of claim 6 further including the step of adjusting the concentration of the collected sparse component according to the relation: ##EQU2## where Q.sub.plasma is the flow rate through the second exit line, Q.sub.collect is the flow rate through the collect line, and Hct.sub.RBC Line is the hematocrit in the first exit line.

### Claim 8 (depends on 7)

The method of claim 7 wherein the rate at which the interface is raised is controlled according to the relation:
Interface Build Rate=(Q.sub.collect +Q.sub.plasma) Hct.sub.RBc Line.
9.

### Claim 9 (depends on 8)

The method of claim 8 further including the steps of
at the conclusion of the spillover phase, providing a system for lowering the level of said interface toward said steady state condition; and
providing a system for repeating the accumulation phase and spillover phase a plurality of times until a desired volume of sparse component is collected.

### Claim 10 (depends on 1)

The method of claim 1 further including the steps of
at the conclusion of the spillover phase, providing a system for lowering the level of said interface toward said steady state condition; and
providing a system for repeating the accumulation phase and spillover phase a plurality of times until a desired volume of sparse component is collected.

### Claim 11 (depends on 1)

The method of claim 1 wherein the step of raising the level of said interface includes adding said more dense component to said vessel at a significant distance from said barrier to minimize disturbance of said pool.

### Claim 12 (depends on 1)

The method of claim 1 wherein the step of establishing the level of said interface in a steady state condition is accomplished by monitoring the level of said interface at a significant distance from said barrier to maintain the level of said more dense component at the monitoring location and allowing the formation of a pool of said sparse component at said barrier.

### Claim 13 (depends on 1)

The method of claim 1 wherein said pool is accumulated by locating said inlet line at a significant distance from said barrier wherein said sparse component is separated from said liquid by centrifugal action by the time the liquid flow reaches the location of said barrier.

### Claim 14 (depends on 1)

The method of claim 1 wherein said liquid is whole blood, said sparse component is essentially white blood cells, said less dense component is essentially plasma, and said more dense component is essentially red blood cells.

### Claim 15 (depends on 14)

The method of claim 14 further including the step of adjusting the concentration of the collected sparse component according to the relation: ##EQU3## where Q.sub.plasma is the flow rate through the second exit line, Q.sub.collect is the flow rate through the collect line, and Hct.sub.RBC Line is the hematocrit in the first exit line.