Apparatus for the sequential performance of chemical processes

• US 4,704,256 A
• Filed: 11/10/1982
• Issued: 11/03/1987
• Est. Priority Date: 09/23/1980
• Status: Expired due to Term

• Current Assignee: Applied Biosystems LLC (Thermo Fisher Scientific Incorporated)
• Original Assignee: California Institute of Technology
• Inventors: Hood, Leroy E., Hunkapiller, Michael W., Hewick, Rodney M., Dreyer, William J., Stark, Anton W.
• Primary Examiner(s): NOT, DEFINED
• Assistant Examiner(s): NOT, DEFINED
• Application Number:
  US06/440,571
• Time in Patent Office: 1,819 Days
• Field of Search: 137/DIG. 8, 210/500.2, 210/282, 239/524, 239/267, 251/61.1, 422/56, 422/58, 422/59, 422/66, 422/68, 422/81, 422/101, 422/103, 422/241, 422/190, 422/211, 422/50, 436/89, 436/177, 436/178, 436/174
• US Class Current: 422/116
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00283 Reactor vessels with top opening
• CPC Class Codes: B01J 2219/00286 Reactor vessels with top and bottom openings
• CPC Class Codes: B01J 2219/00398 in multiple arrangements
• CPC Class Codes: B01J 2219/00418 using pressure
• CPC Class Codes: B01J 2219/00423 using filtration, e.g. through porous frits
• CPC Class Codes: B01J 2219/00495 Means for heating or cooling the reaction vessels
• CPC Class Codes: B01J 2219/0059 Sequential processes
• CPC Class Codes: B01J 2219/00596 Solid-phase processes
• CPC Class Codes: B01J 2219/00686 Automatic
• CPC Class Codes: B01J 2219/00689 using computers
• CPC Class Codes: B01J 2219/00695 Synthesis control routines, e.g. using computer programs
• CPC Class Codes: B01J 2219/00725 Peptides
• CPC Class Codes: B01J 4/02 for feeding measured , i.e. prescribed quantities of reagents
• CPC Class Codes: C40B 40/10 Libraries containing peptides or polypeptides, or derivatives thereof
• CPC Class Codes: C40B 60/14 for creating libraries
• CPC Class Codes: G01N 35/00 Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
• CPC Class Codes: G01N 35/1079 with means for piercing stoppers or septums

Abstract

An improved apparatus and method for the sequential performance of chemical processes on a sample of chemical material wherein the sample is embedded in a solid matrix of fluid permeable material located within a reaction chamber and is sequentially subjected to a plurality of fluids passed through the chamber in a pressurized stream, causing chemical interaction between the sample and the fluids.

Claims

1. A sequenator apparatus for use in the determination of the linear sequence of amino acid units in proteins and peptides, comprising:

chamber means having an interior surface defining a reaction chamber, said chamber having an inlet and an outlet and including a solid, fluid-non-retaining support means within said chamber means;

a plurality of reservoirs containing fluids, at least some of said fluids being interactable with proteins or peptides to perform the degradation process thereof, said reservoirs being in fluid communication with said reaction chamber,

pressure means for moving said fluids from said reservoirs to said reaction chamber,

means for sequentially conducting a plurality of fluids from said reservoirs to said reaction chamber, and through said reaction chamber from said inlet to said outlet in a pressurized stream;

a highly viscous matrix having semisolid properties comprising a thin film on said solid support means which matrix is located within said reaction chamber, said matrix being permeable to said plurality of fluids by diffusion and absorption of a portion of said fluids into said matrix but said matrix being insoluble in said fluids; and

a sample of protein or peptide embedded within said matrix;

whereby said sample is immobilized within said matrix but is exposed to each of the plurality of fluids passing through the reaction chamber for chemical interaction therewith, said exposure being effected by diffusion of a portion of said fluids into the matrix.

2. Apparatus according to claim 1 wherein said matrix comprises a non-reactive polymer.

3. Apparatus according to claim 1 which further comprises means for automatically controlling the functioning of the apparatus through a plurality of cycles.

4. Apparatus according to claim 1 which further includes:

a conversion flask having a plurality of capillary tubes extending into the interior thereof; and

means for introducing various fluids into and withdrawing various fluids from said flask through said capillary tubes, including at least one of said plurality of fluids passed to the outlet of the reaction chamber;

one of said capillary tubes terminating at a point adjacent the bottom of the flask in an inner end at which the bore is closed, said inner end being provided with a plurality of restricted radially spaced orifices;

such that passage of a gas inwardly through said one capillary tube introduces a plurality of small bubbles into any liquid within the flask, said bubbles serving to uniformly agitate or accelerate drying of any liquid within the flask.

5. Apparatus according to claim 1 wherein the sample is a protein or polypeptide which is to be sequentially degraded to determine the sequence of amino acid units therein.

6. Apparatus according to claim 1 which further includes:

a conversion flask having a plurality of capillary tubes extending into the interior thereof; and

means for introducing various fluids into and withdrawing various fluids from said flask through said capillary tubes, including at least one of said plurality of fluids passed to the outlet of the reaction chamber;

at least one of said capillary tubes having an inner end at which the bore is closed and which is provided with a plurality of restricted radially spaced orifices;

such that passage of fluid inwardly through said at least one capillary tube produces a spray onto interior walls of the flask to wash down said interior walls.

7. Apparatus according to claim 6 wherein said capillary tubes comprise glass tubes.

8. Apparatus according to claim 1 wherein said matrix comprises a polymeric quaternary ammonium salt.

9. Apparatus according to claim 8 wherein said polymeric quaternary ammonium salt comprises 1,5-dimethyl-1,5-diazaundecamethylene polymethobromide.

10. Apparatus according to claim 8 wherein said polymeric quaternary ammonium salt comprises poly(N,N-dimethyl-3-5-dimethylene piperidinium chloride).

11. Apparatus according to claim 1 wherein said means for sequentially passing a plurality of fluids through said reaction chamber comprises at least one valve means for controlling the flow of said fluids between the reaction chamber and the plurality of reservoirs, said at least one valve means comprising:

valve block means having a plurality of substantially flat valving sites on a surface thereof, said valve block means defining a plurality passage continuous between two ends thereof and communicating through primary openings with each of said valving sites, and a plurality of secondary passages each communicating through a secondary opening with one of said valving sites;

a plurality of resilient, substantially impermeable diaphragms covering said respective valving sites, each of said diaphragms being actuable between a first sealing position in which it is forced against one of said valving sites to close the primary and secondary openings communicating with said site and a second position in which it is drawn away from said site to provide a fluid flow path between said primary and secondary openings over the exterior of said valve block means; and

means for operatively connecting said primary and secondary passages to said plurality of reservoirs and said reaction chamber;

whereby fluid flow between said plurality of reservoirs and said reaction chamber can be selectively controlled.

12. Apparatus according to claim 11 wherein said connecting means includes a connector fitting at one of said ends of the primary passage such that the primary passage serves as a manifold which can be flushed by a flow of fluid between the connector fitting and a secondary passage furthest away from the connector fitting.

13. Apparatus according to claim 12 wherein said primary passage comprises a plurality of straight passages connected end to end to form a conduit having a sawtooth configuration and communicating at alternating intersections thereof with said respective valving sites.

14. Apparatus according to claim 11 wherein said connecting means comprises at least one tapered ferrule tightly fitted in sliding engagement over a tubing member and urged against a differently tapered recess in said valve block means communicating with one of said passages, such that an inward force applied to said ferrule is focused on a realtively small area of contact between the ferrule and the recess to produce a fluid seal therebetween.

15. Apparatus according to claim 14 wherein said recess is tapered at a greater angle than said ferrule.

16. Apparatus according to claim 14 wherein said ferrule is tapered on opposite sides, one of said sides being urged against said tapered recess by screw thread means having a separate tapered recess for reception of the other side of said ferrule, and recesses being tapered at angles greater than the angles of taper of the corresponding sides of said ferrule.

17. The apparatus of claim 1 wherein said reaction chamber comprises a pair of abutting chamber elements having first and second cavities, respectively, on opposed mating surfaces thereof, said first and second cavities being aligned with each other to form said reaction chamber, said first and second cavities being tapered in directions away from said mating surfaces to locations at which they communicate with said inlet and outlet, respectively.

18. The apparatus of claim 17 wherein said solid support means is a porous sheet positioned within a recess in at least one of said mating surfaces for retention within said reaction chamber in an orientation substantially separating said first and second cavities, said reaction chamber further including a sheet of yielding material located on at least one side of said porous sheet, said sheet of yielding material being sandwiched between said mating surfaces forming a seal therebetween, said sheet of yielding material being permeable to said plurality of fluids.

19. Apparatus according to claim 18 wherein said porous sheet extends substantially transversely across said reaction chamber.

20. Apparatus according to claim 18 wherein said porous sheet is made of a plurality of fibers.

21. Apparatus according to claim 18 wherein said porous sheet is made of a plurality of glass fibers.

22. Apparatus according to claim 18 wherein at least one of said chamber elements includes a raised portion on the mating surface thereof which extends about the cavity therein for comprising said sheet of yielding material against the mating surface of the other chamber element and thus enhancing said seal therebetween.

23. Apparatus according to claim 17 wherein said inlet and outlet comprise a pair of capillary passages extending through said chamber elements and communicating at inner ends with said reaction chamber on opposite sides of the solid support means which solid support means comprises a porous sheet.

24. Apparatus according to claim 23 wherein said capillary passages are coaxial with said reaction chamber and extend therefrom to outer capillary openings in said chamber elements.

25. Apparatus according to claim 23 wherein said chamber elements are of glass and have substantially flat surfaces adjacent said capillary passages.

26. Apparatus according to claim 23 wherein said means for sequentially conducting fluids through the reaction chamber comprises at least one slug of resilient material having a substantially flat sealing end and an axial capillary passage which terminates in an opening at said sealing end, and screw thread means for urging said sealing end axially against one of said capillary passages, such that the axial passage in the slug communicates with said capillary passage and the sealing end of the slug seals against a substantially flat surface of the chamber element adjacent thereto.

27. Apparatus according to claim 26 wherein said at least one slug of resilient material comprises a mass of a fluorocarbon polymer encased in a metallic sleeve.

28. Apparatus according to claim 27 wherein said screw thread means includes a bore for closely receiving said metallic sleeve in an axially sliding relationship to ensure accurate alignment of the passage in the slug with said one capillary passage.