Method for improved focus control in molecular array scanning by using a symmetrical filter to determine in-focus-distance
Automated methods and systems for determining an in-focus-distance for a position on the surface of a molecular array substrate using a molecular array scanner are provided. A signal from a first position of an array substrate is detected and noise is filtered out of the detected signal using a symmetrical filter to produce an in-focus-distance. In one embodiment, the in-focus-distance is utilized as an estimated in-focus-distance at a second position of the array substrate. The method finds use in maintaining the focus of a light source while scanning the array by the scanner. Also provided are methods of assaying a sample using the methods and systems of the invention, and kits for performing the invention. The subject invention finds use in a variety of different applications, including both genomics and proteomics applications.
1. A method of determining an in-focus-distance for a first position on a surface of a biopolymeric array, said method comprising:
detecting a signal from said first position of said surface; and
filtering said signal using a symmetrical filter to determine said in-focus-distance for said first position.
2. The method of claim 1 , wherein said in-focus-distance is used as an estimate of an in-focus-distance of a second position of said biopolymeric array.
3. The method of claim 1 , whereby said symmetrical filter utilizes secondary signals from at least one side-position on each side of said first position to filter said signal from said first position.
4. The method of claim 3 , wherein said first position and said side-positions on each side of said first position are located on a single scan line.
5. The method of claim 4 , wherein said side-positions are arranged in pairs of side-positions that are equidistant from the first position.
6. The method of claim 1 , wherein said symmetrical filter is a zero phase filter.
7. The method of claim 1 , wherein said signal is the sum of a first component signal of an absolute position measuring device and a second component signal of a focus error measuring device.
8. The method of claim 7 , wherein said absolute position measuring device is a quadrature position encoder.
9. The method of claim 7 , wherein said focus error measuring device is a position sensitive detector.
10. The method of claim 1 , wherein said in-focus-distance is within about +/−3–5μm of the focal plane of a focused light.
11. The method of claim 1 , wherein said filtering step removes noise from said signal to allow an accurate determination of said in-focus-distance.
12. The method of claim 11 , wherein said noise is at a frequency of approximately 150–1000 Hz.
13. The method of claim 1 , wherein said signal is a fluorescent signal.
14. The method of claim 1 , wherein the biopolymeric array is an array of polynucleotides or polypeptides.
15. A method of focusing a light source onto a second position on a surface of a biopolymeric array, the method comprising:
(a) predicting an in-focus-distance for a second position on a surface of said biopolymeric array using the method of claim 2 ; and
(b) focusing said light said onto said second position using said predicted in-focus-distance.
16. The method of claim 15 , wherein said focusing step is automatic.
17. The method of claim 16 , wherein said focusing step is performed using a focus control loop.
18. The method of claim 15 , wherein said first position and said second position are on adjacent scan lines.
19. The method of claim 15 , further comprising the step of scanning said second position at said in-focus-distance.
20. The method of claim 19 , wherein said method is employed in a method of maintaining focus of a light source while scanning a biopolymeric array using a scanner system.
21. The method of claim 15 , wherein said scanner is a bidirectional line scanner.
22. A computer-readable medium encoding instructions to direct a machine to perform the method of claim 1 .
23. A biopolymer array scanner comprising a computer-readable medium according to claim 22 .
24. A method of assaying a sample, said method comprising:
(a) contacting said sample with a biopolymeric array of two or more biopolymer ligands immobilized on a surface of a solid support; and
(b) reading said array with a biopolymer array scanner according to claim 23 to obtain a result.
25. The method according to claim 24 , wherein said reading step (b) includes:
(i) symmetrically filtering a detected signal from a first position of a scanned line to determine an estimated in-focus-distance for a second position; and
(ii) automatically focusing a laser of the scanner at said second position using said estimated in-focus-distance.
26. The method according to claim 25 , wherein said biopolymer array is chosen from a polypeptide array and a nucleic acid array.
27. A method comprising transmitting a result obtained from a method of claim 25 , from a first location to a second location.
28. The method of claim 27 , where said second location is a remote location.
29. A method comprising receiving data representing said result of a scan obtained by the method of claim 25 .
30. A kit for use in a biopolymer array optical scanner, said kit comprising:
(a) a computer-readable medium according to claim 22 ; and
(b) instructions for operating said scanner according to said programming.
31. The kit according to claim 30 , wherein said kit further includes at least one reference array.
32. The kit according to claims 31 , wherein the kit further includes instructions for installing the encoded instructions.