Separation of Islets of Langerhans

• US 4,797,213 A
• Filed: 02/24/1987
• Issued: 01/10/1989
• Est. Priority Date: 02/24/1986
• Status: Expired due to Fees

• Current Assignee: Connaught Laboratories Limited
• Original Assignee: Connaught Laboratories Limited
• Inventors: Sun, Anthony M., Parisius, John W.
• Primary Examiner(s): SPEAR JR, FRANK
• Application Number:
  US07/017,844
• Time in Patent Office: 686 Days
• Field of Search: 210/772, 210/768, 210/323.1, 210/335, 210/651, 435/240.21, 435/240.23, 424/104, 30/220, 83/408
• US Class Current: 210/651
• CPC Class Codes: C12M 45/02 by mechanical forces; Stirring; Trituration; Comminuting crushing, pulverizing, disintegrating in general B02C
• CPC Class Codes: C12M 45/09 by enzymatic treatment

Abstract

A semi-automated procedure for the isolation of Islets from the pancreata of different species is described. Pancreatic tissue first is treated to free the Islets from other pancreatic tissue. Enzyme digestion generally is used at least partially to achieve this effect. After separation of the Islet suspension from pancreatic debris, the suspension is subjected to multiple filtration to separate the Islets, which then are recovered from the filter.

Claims

1. A method for the separation of Islets of Langerhans from pancreatic tissue, which comprises:

treating pancreatic tissue to release Islets from other pancreatic tissue to provide an aqueous suspension of Islets and pancreatic tissue debris;

removing said pancreatic tissue debris from said aqueous suspension by filtering off said pancreatic tissue debris and leaving an aqueous suspension of Islets,

removing said Islets from said aqueous suspension by passing said aqueous suspension of Islets successively through a series of screens of decreasing pore size in the direction of flow of said suspension of Islets so as to remove said Islets from said aqueous suspension of Islets on the surface of said screens, and

recovering the Islets from each of said screen surfaces by passing an aqueous wash medium across the surface of each of the screens to remove said Islets from said screen surfaces as an aqueous suspension thereof in said aqueous wash medium.

2. The method of claim 1 wherein said pancreatic tissue is bovine or rat pancreatic tissue and said treatment includes enzymatic digestion.

3. The method of claim 1 wherein said pancreatic tissue is porcine or human pancreatic tissue and said treatment includes maceration.

4. A method of separation of Islets of Langerhans from bovine or rat pancreatic tissue, which comprises:

agitating thin slices of said pancreatic tissue in enzyme solution capable of digestion of pancreatic membrane tissue for a tissue sufficient to release said Islets from said pancreatic tissue, to provide a digest suspension of pancreatic tissue debris Islets,

filtering out said pancreatic tissue debris from said Islet suspension, and

passing the resulting Islet suspension through a series of filter screens of decreasing pore size to filter out Islets from said suspension on said screens to provide an Islet-free digest solution which is recycled to said agitation step, and

washing said Islets from said filter screens by passing in aqueous wash medium across the surface of each screen and collecting the washed Islets.

5. A method of separation of Islets of Langerhans from porcine or human pancreatic tissue, which comprises:

macerating strips of said pancreatic tissue in an aqueous medium for a time sufficient to release said Islets from said pancreatic tissue to provide a suspension of pancreatic tissue debris and Islets,

filtering out said pancreatic tissue debris from said Islet suspension,

passing the resulting Islet suspension through a series of filter screens of decreasing pore size to filter out Islets from said suspension on said screens to provide an Islet-free aqueous medium for discard, and

washing said Islets from said filter screens by passing an aqueous wash medium across the surface of each screen and collecting the washed Islets.

6. A pancreatic cutting tissue apparatus, comprising:

lower horizontal cutting surface means comprising a first plurality of parallel closely-spaced upright thin metal strips mounted in a first frame member, said lower cutting surface means having a plurality of upwardly-opening parallel grooves formed in said first plurality of metal strips and extending transverse to the direction of extension of said strips,

upper cutting surface means comprising a second plurality of closely-spaced upright thin metal strips mounted in a second frame member pivotally connected to said first member and movable to a cutting position wherein said second plurality of metal strips are at least partially interleaved with said first plurality of metal strips, and

means associated with said upper cutting surface for reciprocating said second plurality of metal strips in said second frame member so that said second plurality of metal strips move reciprocably longitudinally relative to said first plurality of metal strips in said cutting position so as to cut pancreatic tissue into thin slices.

7. An apparatus for the separation of Islets of Langerhans from pancreatic tissue, which comprises:

treatment means for releasing Islets from other pancreatic tissue and to provide an aqueous suspension of Islets and pancreatic tissue debris;

first separation means for removing pancreatic tissue debris from said aqueous suspension; and

second separation means for removing Islets from said aqueous suspension, said second separation means comprising filter means, said filter means comprising;

a plurality of parallel filtration compartments separated from one another by filter elements, said filter elements in said compartments having a decreasing pore size from a first filtration compartment to a last filtration compartment,

a first inlet to said first filtration compartment in fluid flow communication with an outlet from said first separation means to receive said aqueous suspension of Islets from said first separation means,

a first outlet from said last filtration compartment so as to establish a path of flow of said Islet suspension from said first inlet to said first outlet and to arrest Islets on the surface of said filter elements, and

a plurality of second inlets and a plurality of second outlets corresponding in number to the number of said compartments and arranged in fluid flow communication with each of said compartments to permit, in such compartment, wash liquid to flow from one of said second inlets to one of said second outlets across the surface of the filter element to wash filtered Islets therefrom and out of the filter means through the corresponding one of said second outlets.

8. The apparatus of claim 7 wherein said treatment means comprises a digestion vessel having a first inlet for digestion enzyme solution, a second inlet for sliced pancreatic tissue to be digested and an outlet for liquid digest, and said first separation means comprises a screen positioned in said vessel to retain partially-digested pancreatic tissue while permitting freed Islets to pass out of the digestion vessel through the outlet.

9. The apparatus of claim 7 wherein said treatment means comprises a macerator having an upper inlet for roughly chopped pancreatic tissue, a plurality of pairs of intermeshing grooved rollers arranged in a vertical stack, with the grooves progressing from coarser to a finer from top to bottom and a lower outlet for separated Islets, and said first separation means comprises a screen positioned in said vessel to retain pancreatic tissue debris while permitting freed Islets to pass out of the macerator through the outlet