Automated apparatus for pancreatic islet isolation and processing

• US 8,030,058 B1
• Filed: 03/03/2006
• Issued: 10/04/2011
• Est. Priority Date: 01/23/2001
• Status: Expired due to Fees

• Current Assignee: Daniel J. Benedict, Lorna S. Mosse
• Original Assignee: Daniel J. Benedict, Lorna S. Mosse
• Inventors: Benedict, Daniel J., Mosse, Lorna S.
• Primary Examiner(s): Marcheschi; Michael
• Assistant Examiner(s): MA, JAMESON Q
• Application Number:
  US11/367,188
• Time in Patent Office: 2,041 Days
• Field of Search: 4352861-2867
• US Class Current: 435/286.5
• CPC Class Codes: C12M 47/04 Cell isolation or sorting purging biological preparations of unwanted cells C12N5/0081, determining the presence or kind of microorganism C12Q1/04

Abstract

Advanced islet separation technology incorporates an automated method, automated control methodology, process control interface, and automated apparatus to separate (isolate) and process pancreatic islets in a tissue suspension in physiologic process solution, utilizing microprocessor control and/or microprocessor computer control and software programming (code) to interface and control the process temperature, fluid flow rate, pH, dissolved oxygen concentration, endotoxin concentration, dissolved nitric oxide concentration, nitric oxide synthase activity, proteolytic enzyme activity, and/or pressure of the islet containing physiologic process solution, the extent of islet separation, including real-time data acquisition and recording of process variables.

Claims

1. An apparatus for isolating islets from a pancreatic tissue, comprising:

a heated feed tank;

a reactor for containing a physiologic process solution having a dissolved oxygen concentration, an antibiotic concentration, a pH, a temperature, and a flowrate said reactor comprising a flow loop that connects a digestion chamber, a sparging vessel and a process pump, said flow loop being connected through a first valve to said heated feed tank, said process pump being operative to circulate said physiologic process solution through said reactor;

a process controller;

a sensor block that is located on said flow loop upstream from said digestion chamber, said sensor block having a plurality of sensors that send signals to said process controller, said sensor block comprising a dissolved oxygen sensor for sensing said dissolved oxygen concentration, an antibiotic sensor for sensing said antibiotic concentration, a pH sensor for sensing said pH, a temperature sensor for sensing said temperature, and a flow sensor for sensing said flowrate;

a source of oxygen gas controlled by a oxygen gas valve for introducing a stream of oxygen gas into said sparging vessel, a source of antibiotic controlled by a pump for introducing antibiotic into the process, and a source of an inert gas controlled by an inert gas valve for introducing a stream of inert gas into said sparging vessel;

an auto-collector that is connected to said flow loop for collecting the islets;

an integrated flow-through sample cell for monitoring the extent of pancreatic digestion and islet separation in real-time;

a proteolytic process controller which deactivates proteolytic enzyme activity and ends digestion by adding an effective amount of an antibiotic in response to a signal derived from said flow through sample cell; and

wherein said process controller compares the dissolved oxygen concentration in said physiologic process solution to a dissolved oxygen setpoint and activate either said oxygen gas valve to add dissolved oxygen or said inert gas valve to remove dissolved oxygen, to compare said antibiotic concentration setpoint and activate control of antibiotic concentration, compares said pH to a pH setpoint and activate either said acid solution pump to reduce said pH or said base solution pump to increase said pH, compares said temperature to a temperature setpoint and to activate either said process cooler to decrease said temperature or said process heater to increase said temperature, and compares said flowrate to a flowrate setpoint and control said flowrate.

2. The apparatus of claim 1 wherein said plurality of solution pumps further comprise a dissolved nitric oxide control means selected from the group consisting of an antibiotic solution pump for pumping an antibiotic solution from an antibiotic solution reservoir into said flow loop, a nitric oxide scavenger solution pump for pumping a nitric oxide scavenging solution from a nitric oxide scavenger solution reservoir into said flow loop, and an amino acid solution pump for pumping an amino acid solution from an amino acid solution reservoir into said flow loop;

and wherein said process controller is operative to compare said dissolved nitric oxide concentration to a dissolved nitric oxide concentration set point and activate said antibiotic solution pump, said nitric oxide scavenger solution pump or said amino acid solution pump.

3. The apparatus of claim 1 wherein moving means are provided that are operative to rotate, move linearly or move eccentrically said digestion chamber, valve means are provided that are operative to circulate the physiologic process solution through said digestion chamber in a forward direction and in a reverse direction and transducer means are provided that are operative to cause sonication of the physiologic process solution circulating through said digestion chamber.

4. The apparatus of claim 1, wherein the opening and closing of said valves is controlled by manual manipulation of said computer by a user.

5. The apparatus of claim 1, wherein said computer further comprises a graphical user interface to facilitate manual manipulation of said computer by said user.

6. The apparatus of claim 1, further comprising a flow through sample cell to monitor and control the extent of islet separation via image analysis.

7. The apparatus of claim 1 wherein said physiologic process solution further has an endotoxin concentration;

wherein said sensor block further comprises an endotoxin sensor assembly for sensing said endotoxin concentration;

wherein said plurality of solution pumps further comprise an endotoxin neutralizing solution pump for pumping an endotoxin neutralizing solution from an endotoxin neutralizing solution reservoir into said flow loop; and

wherein said process controller is operative to compare said endotoxin concentration to an endotoxin concentration set point and activate said endotoxin neutralizing solution pump.

8. The apparatus of claim 2 wherein nitric acid control comprises an inert gas valve.