CN-121994685-A - Flow cytometer with variable depth sample line injection tube and method of use thereof

CN121994685ACN 121994685 ACN121994685 ACN 121994685ACN-121994685-A

Abstract

Aspects of the present disclosure include flow cytometers having variable depth sample line injection tubes. A flow cytometer according to certain embodiments includes a flow cell, and a Sample Injection Tube (SIT) assembly including a sample line operably coupled to the flow cell, wherein the SIT assembly is configured to provide a variable sample line depth in a sample container. Methods of treating samples with flow cytometry in, for example, analytical and/or sorting applications are also provided.

Inventors

K. T. Dembsky
H. J. Rankila
R.G.Fang

Assignees

贝克顿·迪金森公司

Dates

Publication Date: 20260508
Application Date: 20251028
Priority Date: 20241108

Claims (15)

1. A flow cytometer, comprising: A flow cell; A sample container receiving area; A Sample Injection Tube (SIT) assembly comprising a sample line fluidly coupled to the flow cell, and An actuator coupled with the SIT component, the actuator configured to move the SIT component; Wherein the SIT assembly is configured to provide a variable sample line depth in a sample container present in the sample container receiving area.
2. The flow cytometer of claim 1, wherein the SIT component comprises: An arm having a first proximal end coupled to the actuator and a second distal end including a through bore, and A sample line subassembly located in the through-hole; Wherein the sample line subassembly is reversibly movable in a z-direction relative to an x-y plane of the second end.
3. The flow cytometer of claim 2, wherein the sample line subassembly is reversibly movable in a z-direction by a predetermined displacement distance relative to an x-y plane of the second end.
4. A flow cytometer according to claim 3 wherein the predetermined displacement distance is in the range of 1 mm to 2 mm.
5. The flow cytometer of claim 4, wherein the predetermined displacement distance range is 1.5 mm.
6. The flow cytometer of any of claims 2 to 5, wherein the sample line subassembly comprises: a threaded bushing, at least a portion of which is present in the through bore, wherein the threaded bushing includes a threaded bore and a sample line bore passing through the threaded bore, and A sample line screw coupled with the threaded bore and including an internal bore receiving the sample line.
7. The flow cytometer of claim 6, wherein the sample line subassembly further comprises a ferrule in the threaded bore, wherein the sample line passes through the ferrule.
8. The flow cytometer of any of claims 2 to 7, wherein the SIT assembly further comprises a spring configured to press the sample line subassembly onto the arm.
9. The flow cytometer of claim 8, wherein the spring comprises a return flexure.
10. The flow cytometer of claim 9, wherein the return flexure comprises an opening at a first end through which the sample line screw of the sample line subassembly passes.
11. The flow cytometer of claim 10, wherein the return flexure is coupled with the arm.
12. The flow cytometer of any of claims 2 to 11, wherein the SIT assembly further comprises a stop member configured to limit movement of the sample line subassembly in the z direction relative to the x-y plane of the second end.
13. The flow cytometer of claim 12, wherein the flow cytometer further comprises a sensor that measures the displacement of the return flexure.
14. The flow cytometer of claim 13, wherein the flow cytometer is configured to evaluate sample line positioning in a sample container based on the output of the sensor.
15. A method of analyzing a sample, the method comprising: (a) Introducing a particle sample from a sample container into a flow cytometer with a Sample Injection Tube (SIT) assembly comprising a sample line operably coupled to a flow cell, wherein the SIT assembly is configured to provide a variable sample line depth in the sample container, and (B) The samples were analyzed by flow cytometry.

Description

Flow cytometer with variable depth sample line injection tube and method of use thereof Cross Reference to Related Applications According to 35 U.S. c. ≡119 (e), the present application claims priority from the filing date of U.S. provisional patent application serial No. 63/718,451 filed on 8 th 11 of 2024, the disclosure of which is incorporated herein by reference in its entirety. Background Characterization of analytes in biological fluids has become an important part of biological research, medical diagnostics, and overall health assessment of patients. Detection of analytes in biological fluids (e.g., human blood or blood-derived products) may provide results that may play a role in determining treatment regimens for patients suffering from various conditions. Flow cytometry is a technique for characterizing and often sorting biological materials, such as cells of a blood sample or particles of interest in another biological or chemical sample. Flow cytometers typically include a sample reservoir for receiving a fluid sample (e.g., a blood sample) and a sheath fluid reservoir containing a sheath fluid. The flow cytometer delivers particles (including cells) in a fluid sample as a stream of cells to a flow cell while also directing sheath fluid to the flow cell. To characterize the composition of the flow stream, the flow stream is irradiated with light. Changes in the material in the flow stream (e.g., the presence of morphological or fluorescent markers) can result in changes in the observed light, and these changes can be used for characterization and separation. To characterize the composition in a flow stream, light must impinge on the flow stream and be collected. The light source in the flow cytometer may vary and may include one or more of a broad spectrum lamp, a light emitting diode, and a single wavelength laser. The light source is directed at the flow stream and collects and quantifies the optical response from the irradiated particles. Separation of biological particles is achieved by adding a sorting or collection function to the flow cytometer. Particles in the separation stream that are detected as having one or more desired characteristics are separated from the sample stream by mechanical or electrical removal. One common flow sorting technique utilizes droplet sorting in which a flowing stream containing linear separator particles is broken up into droplets. Droplets containing the particles of interest are charged and deflected into a collection tube by passing through an electric field. Typically, linear spacer particles in the stream are characterized as they pass through a viewpoint located directly below the nozzle tip. Once it is determined that a particle meets one or more desired criteria, it can be predicted when it will reach the drop separation point and separate from the stream in the form of a drop. Ideally, the flow stream is subjected to a transient charge immediately prior to separation of the droplets containing the selected particles from the flow stream, and then grounded immediately after separation of the droplets. The droplets to be sorted remain charged when they are detached from the flow stream, while all other droplets are uncharged. The sample analyzed in the flow cytometer may initially be placed in a sample container, such as a tube or well of a multi-well plate. For introducing the sample from the sample container into the flow cell of the flow cytometer, a sample injection tube may be employed. FIG. 1A provides a view of a sample injection tube commonly found in currently used flow cytometers. As shown in fig. 1A, sample injection tube 10 includes a sample line 25 secured to a first end 40 of support arm 30 by a sample line screw 15. Since the sample line screw secures the sample line to the end 40 of the support arm 30, the sample line 25 does not move in the z-direction relative to the end 40 of the support arm 30. Also shown is an outer sleeve 20 as part of the droplet containment system. The support arm 30 is coupled at a distal end 45 to a first actuator 35 that moves the support arm 30 in the z-direction relative to the outer sleeve 20 so that the distal end 25a of the sample line 25 can be moved into the outer sleeve 20 for washing or the outer sleeve can be moved out to withdraw a sample from a sample container. Also shown is a second actuator 50 driven by a motor to move the entire assembly up and down in the z-direction. Disclosure of Invention The inventors realized that the sample injection tube designs currently in use (as shown in fig. 1A) do not contemplate different sample container bottom configurations. Thus, the sample injection tube configurations currently in use can result in varying amounts of dead volume (i.e., sample in the container that is not aspirated into the flow cytometer) that can vary from sample container type to sample container type and manufacturer. The dead volume may vary in a given workflow, and in some cases ra