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CN-121994682-A - Flow cytometer with pneumatically driven autosampler and method of use thereof

CN121994682ACN 121994682 ACN121994682 ACN 121994682ACN-121994682-A

Abstract

Aspects of the present disclosure include flow cytometers having pneumatically driven autosamplers. According to some embodiments, a flow cytometer includes a flow cell and a pneumatically driven autosampler configured to automatically obtain a sample from a sample container and deliver the sample to the flow cell. The present disclosure also provides methods of flow cell processing of samples, for example, in analytical and/or sorting applications.

Inventors

  • H. J. Rankila
  • J. Manzalaga
  • J. Fendberg

Assignees

  • 贝克顿·迪金森公司

Dates

Publication Date
20260508
Application Date
20251127
Priority Date
20241108

Claims (15)

  1. 1. A flow cytometer, comprising: A flow chamber, and A pneumatically driven autosampler configured to automatically take a sample from a sample container and deliver the sample to the flow chamber.
  2. 2. The flow cytometer of claim 1 wherein the pneumatically driven autosampler comprises: A sample container receiving area; a SIT assembly configured to introduce a sample line into the sample container in the sample container region, wherein SIT refers to a sample injection tube, and A sample gate configured to regulate access to the sample container receiving area; Wherein actuation of the SIT assembly and the sample gate is pneumatically driven by a pneumatic assembly.
  3. 3. The flow cytometer of claim 2 wherein the pneumatic assembly comprises: A pneumatic pump for providing positive pressure to the first pneumatic line; A first switch in fluid communication with the pneumatic pump via the first pneumatic line and configured to direct positive pressure to a second pneumatic line and a third pneumatic line such that when positive pressure is applied to the second pneumatic line, the third pneumatic line is not pressurized, and vice versa; a first cylinder in fluid communication with the second pneumatic line and the third pneumatic line and mechanically coupled to the SIT assembly such that pressurization of the second pneumatic line moves the SIT assembly to a sampling position; a second switch in fluid communication with the pneumatic pump via the first pneumatic line and configured to direct positive pressure to a fourth pneumatic line and a fifth pneumatic line such that when positive pressure is applied to the fourth pneumatic line, the fifth pneumatic line is not pressurized, and vice versa; A second cylinder in fluid communication with the fourth pneumatic line and the fifth pneumatic line and in gate-actuated connection with the sample gate such that pressurization of the fourth pneumatic line moves the sample gate to a closed position and pressurization of the third pneumatic line moves the sample gate to an open position.
  4. 4. The flow cytometer of claim 3 wherein the pneumatic assembly further comprises: A third switch in fluid communication with the pneumatic pump via the first pneumatic line and configured to direct positive pressure to a sixth pneumatic line and a seventh pneumatic line such that when positive pressure is applied to the sixth pneumatic line, the seventh pneumatic line is not pressurized, and vice versa; A third cylinder in fluid communication with the sixth pneumatic line and the seventh pneumatic line and mechanically coupled to a sampling tube subassembly, the sampling tube subassembly including a sample line in fluid communication with the flow chamber such that pressurization of the sixth pneumatic line moves the sample line subassembly to a sample introduction position, thereby causing sample line insertion into a sample container, and pressurization of the seventh pneumatic line moves the sample tube subassembly to a retracted position.
  5. 5. The flow cytometer of claim 3 or 4 wherein actuation of the SIT assembly, the sample gate, and/or the sample tube subassembly is not driven by a stepper motor and worm gear.
  6. 6. The flow cytometer of any of claims 3 to 5 wherein actuation of the SIT assembly, the sample gate, and/or the sample tube subassembly does not require a separate circuit board.
  7. 7. The flow cytometer of any of claims 3 to 6 wherein the pneumatic pump, first, second, and third cylinders, and first, second, and third switches are operated by a single circuit board.
  8. 8. The flow cytometer of any of claims 3 to 7 wherein the pneumatic pump, first, second, and third cylinders, and first, second, and third switches do not require firmware.
  9. 9. The flow cytometer of any of claims 3 to 8 wherein one or more of the first, second, and third switches comprises a flow regulator for regulating pressure balance in a non-pressurized pneumatic line.
  10. 10. The flow cytometer of any of claims 3 to 9 wherein the pneumatic assembly further comprises a pressure reservoir in fluid communication with the pneumatic pump and one or more of the first, second, and third switches, wherein the pneumatic pump pressurizes the pressure reservoir and the pressure reservoir provides positive pressure to the first pneumatic line.
  11. 11. The flow cytometer of any of claims 3 to 10 wherein the pneumatic assembly comprises a connector on one or more of the first, second, third, fourth, fifth, sixth, and seventh pneumatic lines such that the pneumatic lines can be disconnected and reconnected.
  12. 12. The flow cytometer of any of claims 3 to 11 wherein the pneumatic assembly further comprises a mounting bracket on which the pneumatic pump, pressure reservoir, switch, and connector are mounted.
  13. 13. The flow cytometer of any of claims 3 to 12, the pneumatic assembly further comprising a pressure gauge for measuring pressure within the pressure reservoir and one or more of the second, third, fourth, fifth, sixth, and seventh pneumatic lines.
  14. 14. The flow cytometer of any of claims 2 to 13 wherein the sample gate protects the sample from ambient light by reducing transmission of ambient light.
  15. 15. A method of flow cytometry analysis of a sample, the method comprising: (a) Introducing a sample container containing a sample into a pneumatically driven autosampler of the flow cytometer, wherein the pneumatically driven autosampler is configured to automatically obtain a sample from the sample container, thereby delivering the sample to a flow cell of the flow cytometer; (b) And performing flow cytometry analysis on the sample.

Description

Flow cytometer with pneumatically driven autosampler and method of use thereof Cross Reference to Related Applications The present application claims priority to the date of filing of U.S. provisional application No. 63/813,208 filed on month 28 of 2025 and U.S. provisional application No. 63/718,455 filed on month 8 of 2024 in accordance with the provisions of chapter 35, section 119 (e) of the united states code of america, the disclosures of which are incorporated herein by reference. Background Characterization of analytes in biological fluids has become an important component of biological research, medical diagnosis, and assessment of overall patient health and wellbeing. Detection of analytes in biological fluids, such as human blood or blood-derived products, may provide results that may be useful in determining treatment regimens for patients suffering from a variety of disease states. Flow cytometry is a technique for characterizing and often sorting biological materials, such as cells in a blood sample or other types of biological or chemical samples, of particles of interest. Flow cytometers typically include a sample reservoir for receiving a fluid sample, such as a blood sample, and a sheath fluid reservoir containing a sheath fluid. The flow cytometer delivers particles (including cells) in the fluid sample as a stream of cells to the 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, such as the presence of morphological or fluorescent markers, may result in changes in the observed light, and these changes can be used for characterization and separation. In order to characterize the components in the flow stream, light must be directed onto the flow stream and collected. The light source of 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 aligned with the flow stream and the optical response from the illuminated particles is collected and quantified. The separation of biological particles can be achieved by adding a sorting or collecting function to the flow cytometer. In the separated stream, particles detected to have one or more desired characteristics are separated from the sample stream by mechanical or electrical removal. One common flow sorting technique employs droplet sorting, in which a stream containing linearly separated particles is broken up into droplets. Droplets containing the particles of interest are charged and deflected into a collection tube when passing through an electric field. Typically, particles that are linearly separated in the stream are characterized as they pass through a viewpoint located directly below the tip of the nozzle. Once a particle is identified as meeting one or more desired criteria, it can be predicted when it will reach the point of droplet break and separate into droplets from the stream. Ideally, the convection current is briefly charged immediately before the drop containing the selected particle breaks away from the fluid stream, and then grounded immediately after the drop breaks away. The drop to be sorted remains charged while it is detached from the fluid flow, while all other drops remain uncharged. The sample analyzed in the flow cytometer may initially be contained 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 used. FIG. 1A provides a sample injection tube common in currently used flow cytometers. As shown in fig. 1A, sample injection tube assembly 10 includes a sample tube 25 secured to a first end 40 of a support arm 30 by a sample tube bolt 15. Since the sample line bolts secure the sample line at 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 that is part of the droplet containment system. One end 45 of the support arm 30 is connected to a first actuator 35 that moves the support arm in the z-direction relative to the outer cannula 20 so that the distal end 25a of the sample line 25 can be moved to the outer cannula 20 for washing or can be extended from the outer cannula 20 to aspirate a sample from a sample container. Also shown is a second actuator 50, which is a motor driven actuator, that moves the entire assembly up and down in the z-direction. Disclosure of Invention The inventors have recognized that a motor driven actuator as shown in fig. 1A requires additional Printed Circuit Boards (PCBs) and firmware to operate the motor in addition to the motor and gears. Therefore, the structure of these devices is very complicated, resulting in an increase in manufacturing costs and maintenance costs. In addition, th