Microfabricated two-pin system for biomolecule crystallization

• US 7,153,699 B2
• Filed: 12/23/2002
• Issued: 12/26/2006
• Est. Priority Date: 12/21/2001
• Status: Expired due to Term

• Current Assignee: Cytonome/St, Llc (Cytonome Inc.)
• Original Assignee: Cytonome Inc.
• Inventors: Gilbert, John, Trikha, Jaishree
• Primary Examiner(s): GORDON, BRIAN R
• Application Number:
  US10/328,932
• Publication Number:
  US 20030170146A1
• Time in Patent Office: 1,464 Days
• Field of Search: 436/174, 436/180, 422/100, 738/647.2
• US Class Current: 436/180
• CPC Class Codes: B01L 2200/142 Preventing evaporation
• CPC Class Codes: B01L 2200/143 Quality control, feedback systems
• CPC Class Codes: B01L 2200/147 Employing temperature sensors
• CPC Class Codes: B01L 2300/1822 using Peltier elements
• CPC Class Codes: B01L 2400/0439 ultrasonic vibrations, vibrating piezo elements
• CPC Class Codes: B01L 3/0244 using pins
• CPC Class Codes: B01L 3/0248 Prongs, quill pen type dispenser
• CPC Class Codes: B01L 3/0255 characterized by the form or material of the pin tip
• CPC Class Codes: B01L 3/06 Crystallising dishes
• CPC Class Codes: B01L 3/5088 confining liquids at a location by surface tension, e.g. virtual wells on plates, wires B01L3/50857 takes precedence
• CPC Class Codes: C40B 40/10 Libraries containing peptides or polypeptides, or derivatives thereof
• CPC Class Codes: C40B 50/08 Liquid phase synthesis, i.e. wherein all library building blocks are in liquid phase or in solution during library creation; Particular methods of cleavage from the liquid support
• CPC Class Codes: G01N 2035/1037 Using surface tension, e.g. pins or wires
• CPC Class Codes: Y10T 436/25 including sample preparation
• CPC Class Codes: Y10T 436/2575 Volumetric liquid transfer

Abstract

A two-pin liquid handling system for crystallizing biomolecules comprises a pair of interacting pins for holding a droplet of liquid therebetween. Each pin includes a tip spaced predetermined distance from the other pin to define a sample acquisition region. The two-pin liquid handling system includes a temperature and humidity control element for controlling the temperature of the sample acquisition region to cause transpiration of water to and from a droplet of biomolecule solution to promote crystallization of the biomolecule.

Claims

1. A method for crystallizing biomolecules, comprising the steps of:

providing a liquid handling assembly comprising a plurality of components formed on a single substrate for handling a fluid, the plurality of components comprising a holder, a first pin extending from and integral with the holder, a second pin extending from and integral with the holder, the second pin spaced from the first pin to define a channel therebetween sized and dimensioned for receiving and retaining a fluid sample by capillary action;

acquiring a droplet of a biomolecule solution in the channel;

transpiring water between the biomolecule solution and the environment to concentrate the biomolecule, thereby causing crystallization of the biomolecule to occur within the channel.

2. The method of claim 1, wherein the channel has a capacity of between about 0.1 nanoliters and about 50 nanometers.

3. The method of claim 1, further comprising the step of:

observing formation of a crystal.

4. The method of claim 3, wherein the step of observing comprises detecting biomolecule aggregation by laser scattering.

5. The method of claim 3, wherein the step of observing comprises using an optical device selected from the group consisting of a polarization microscope, a CCD camera, and a video camera.

6. The method of claim 1, further comprising the step of freezing the crystal on the pin.

7. The method of claim 1, wherein the step of acquiring a droplet of biomolecule solution comprises dipping the pin in a well containing a supply of the biomolecule solution and a crystallization reagent.

8. The method of claim 1, wherein the step of transpiring comprises controlling the temperature of one of the first pin and the second pin.

9. The method of claim 1, wherein the first pin and second pin are separated by a distance of less than about 100 microns where the channel is formed.

10. The method of claim 1, wherein the first pin has a first tip having a concavely curved surface and the second pin has a second tip with a concavely curved surface spaced a predetermined separation distance from the first tip to define the channel between the concavely curved surfaces.

11. The method of claim 10, wherein the concave surface of the first tip faces and the predetermined separation distance between the two concave surfaces is less than about 100 microns.

12. The method of claim 1, wherein each pin has a tip defining a side of the channel, and each tip comprises a body connecting the tips to the holder, and the body of the first pin is separated from the body of the second tip by a second distance that is larger than the predetermined separation distance between the first tip and second tip.

13. The method of claim 8, wherein the step of controlling the temperature employs a Peltier element.

14. The method of claim 3, further comprising the step of altering crystallization conditions in response to the step of observing.

15. The method of claim 14, wherein the step of altering comprising varying the temperature of the crystallizing structure.

16. The method of claim 14, further comprising the steps of:

recording the crystallization conditions; and

observing the crystal formed in the channel.

17. The method of claim 8, wherein the liquid handling system further comprises a temperature control element, a temperature sensor for sensing the temperature of the system, and a controller in circuit with the temperature control element and the temperature sensor for controlling the temperature of the liquid handling system.

18. The method of claim 8, wherein the substrate comprises a silicon wafer.