Automated sorting system for matrices with memory

• US 6,329,139 B1
• Filed: 08/11/1997
• Issued: 12/11/2001
• Est. Priority Date: 04/25/1995
• Status: Expired due to Fees

• Current Assignee: Irori Technologies Incorporated
• Original Assignee: Discovery Partners International, Inc. (Infinity Pharmaceuticals, Inc.)
• Inventors: Lillig, John E., Ewing, William, Karunaratne, Kanchana Sanjaya Gunesekera, Nova, Michael P., Potash, Hanan, Satoda, Yozo
• Primary Examiner(s): Horlick, Kenneth R.
• Assistant Examiner(s): Siew, Jeffrey
• Application Number:
  US08/912,998
• Time in Patent Office: 1,583 Days
• Field of Search: 209/597, 209/604, 364/130, 435/6
• US Class Current: 506/30
• 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 2219/00295 the reactor vessels having pervious side walls
• CPC Class Codes: B01J 2219/00299 Generally cylindrical reactor vessels
• CPC Class Codes: B01J 2219/00308 interchangeably mounted in racks or blocks
• CPC Class Codes: B01J 2219/0031 the racks or blocks being mounted in stacked arrangements
• CPC Class Codes: B01J 2219/00313 the reactor vessels being formed by arrays of wells in blocks
• CPC Class Codes: B01J 2219/00319 the blocks being mounted in stacked arrangements
• CPC Class Codes: B01J 2219/00333 Closures attached to the reactor vessels
• CPC Class Codes: B01J 2219/00344 Caps
• CPC Class Codes: B01J 2219/00454 by chemical cleavage from the solid support
• CPC Class Codes: B01J 2219/00461 Beads and reaction vessel together
• CPC Class Codes: B01J 2219/00463 Directed sorting
• CPC Class Codes: B01J 2219/005 Beads
• CPC Class Codes: B01J 2219/00502 Particles of irregular geometry
• CPC Class Codes: B01J 2219/0054 Means for coding or tagging the apparatus or the reagents
• CPC Class Codes: B01J 2219/00542 Alphanumeric characters
• CPC Class Codes: B01J 2219/00547 Bar codes
• CPC Class Codes: B01J 2219/00549 2-dimensional
• CPC Class Codes: B01J 2219/00551 3-dimensional
• CPC Class Codes: B01J 2219/00554 : Physical means
• CPC Class Codes: B01J 2219/0056 : Raised or sunken areas
• CPC Class Codes: B01J 2219/00563 : Magnetic means
• CPC Class Codes: B01J 2219/00567 : Transponder chips
• CPC Class Codes: B01J 2219/00569 : EEPROM memory devices
• CPC Class Codes: B01J 2219/00585 : Parallel processes
• CPC Class Codes: B01J 2219/0059 : Sequential processes
• CPC Class Codes: B01J 2219/00592 : Split-and-pool, mix-and-divide processes
• CPC Class Codes: B01J 2219/00596 : Solid-phase processes
• CPC Class Codes: B01J 2219/00689 : using computers
• CPC Class Codes: B01J 2219/00691 : using robots
• CPC Class Codes: B01J 2219/00695 : Synthesis control routines, e.g. using computer programs
• CPC Class Codes: B01J 2219/0072 : Organic compounds
• CPC Class Codes: B01J 2219/00722 : Nucleotides
• CPC Class Codes: B01J 2219/00725 : Peptides
• CPC Class Codes: B01L 2300/022 : Transponder chips
• CPC Class Codes: B01L 2300/0609 : Holders integrated in container to position an object
• CPC Class Codes: B01L 2400/0655 : pinch valves
• CPC Class Codes: B01L 3/50255 : Multi-well filtration
• CPC Class Codes: B01L 3/50825 : Closing or opening means, corks, bungs closures for containers B65D; means for removing stoppers B67B7/02
• CPC Class Codes: B01L 3/5453 : for test tubes
• CPC Class Codes: B82Y 10/00 : Nanotechnology for information processing, storage or transmission, e.g. quantum computing or single electron logic
• CPC Class Codes: C07H 21/00 : Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C07K 1/00 : General methods for the preparation of peptides , i.e. processes for the organic chemical preparation of peptides or proteins of any length
• CPC Class Codes: C07K 1/04 : on carriers C07K1/003, C07K1/006 take precedence
• CPC Class Codes: C07K 1/047 : Simultaneous synthesis of different peptide species; Peptide libraries
• CPC Class Codes: C08J 2327/12 : containing fluorine atoms
• CPC Class Codes: C08J 7/16 : with polymerisable compounds
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 40/10 : Libraries containing peptides or polypeptides, or derivatives thereof
• CPC Class Codes: C40B 50/14 : Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support
• CPC Class Codes: C40B 60/14 : for creating libraries
• CPC Class Codes: C40B 70/00 : Tags or labels specially adapted for combinatorial chemistry or libraries, e.g. fluorescent tags or bar codes
• CPC Class Codes: G01N 15/149 : specially adapted for sorting particles, e.g. by their size or optical properties
• CPC Class Codes: G01N 2035/00782 : reprogrammmable code
• CPC Class Codes: G01N 2333/9121 : with an alcohol group as acceptor (2.7.1), e.g. general tyrosine, serine or threonine kinases
• CPC Class Codes: G01N 2500/00 : Screening for compounds of potential therapeutic value
• CPC Class Codes: G01N 33/542 : with steric inhibition or signal modification, e.g. fluorescent quenching
• CPC Class Codes: G01N 33/54353 : with ligand attached to the carrier via a chemical coupling agent coatings G01N33/54393
• CPC Class Codes: G01N 33/54393 : Improving reaction conditions or stability, e.g. by coating or irradiation of surface, by reduction of non-specific binding, by promotion of specific binding
• CPC Class Codes: G01N 35/00732 : Identification of carriers, materials or components in automatic analysers
• CPC Class Codes: G01N 35/00871 : Communications between instruments or with remote terminals
• CPC Class Codes: G11C 13/0014 : comprising cells based on organic memory material
• CPC Class Codes: G11C 13/0019 : comprising bio-molecules
• CPC Class Codes: G11C 13/025 : using fullerenes, e.g. C60, or nanotubes, e.g. carbon or silicon nanotubes
• CPC Class Codes: G11C 2213/81 : Array wherein the array conductors, e.g. word lines, bit lines, are made of nanowires

Abstract

Automated drug discovery protocols, or partially automated protocols, in which matrices with memories serve as the platform on which all manipulations are performed or serve as the repository of information that is transferred to other memories as the synthesized compounds are processed and screened. Also provided are automated drug discovery units for use in the protocols. The units provide a means for seamless data tracking and include instrumentation and vials with memories for information transfer to other memories in a unit. The units, which are provided herein, include some or all of the following: an automated or manual sorter, microvessels, which contain memories, an automated or semi-automated synthesizer, a microvessel washer/dryer, a manual or automated cleaver for removing compounds from the matrix with memory microvessels, and associated software. The memories may be any of any type, including electromagnetically encodable memories and optical memories, or combinations thereof. The memories may be pre-encoded or may be encodabie during, after or before processing. Also provided are manual and automated methods for sorting matrices with memories.

Claims

1. A system for use in solid phase synthesis comprising:

a plurality of combinations of matrices with memories, each combination of matrix with memory comprising a combination of a matrix material for supporting solid phase synthesis and a remotely-accessible memory containing encoded data for uniquely identifying each matrix with memory;

an identification station for identifying each matrix with memory when the matrix with memory is disposed within a reading range of the identification station;

a plurality of synthesis containers, each synthesis container adapted for receiving one or more selected matrices with memories, each synthesis container containing a solution comprising a building block for solid phase synthesis on the selected matrices with memories;

a host computer in electrical communication with the identification station and having a database comprising a plurality of synthesis sequences, each synthesis sequence comprising a plurality of steps for synthesizing a molecule or biological particle from a plurality of building blocks and, each synthesis sequence corresponding to an identified matrix with memory, wherein the host computer determines a destination of the identified matrix with memory, the destination comprising a pre-determined synthesis container of the plurality of synthesis containers; and

a positioning means responsive to the host computer for moving the matrix with memory to the predetermined destination.

2. The system of claim 1, further comprising means for generating a cue that identifies the pre-determined synthesis container to the positioning means.

3. The system of claim 2, wherein the means for generating the cue is removably mounted on the destination container.

4. The system of claim 1, wherein the remotely-accessible memory comprises an optically-readable symbol and the identification station comprises:

a light emitter and a light detector, wherein by placing a matrix with memory near the light emitter, light is reflected from the optically-readable symbol to the light detector which decodes the reflected light and yields the unique identity of the matrix with memory.

5. The system of claim 1, wherein the remotely-accessible memory comprises an electronic memory connected to a transceiver and the identification station comprises:

an electromagnetic transmitter for generating an electromagnetic field;

and an electromagnetic receiver, wherein placing a matrix with memory within the electromagnetic field activates the memory which in turn transmits information included within the memory to the electromagnetic receiver.

6. The system of claim 1, further comprising a data base within the host computer having a plurality of data fields comprising records of the matrices with memories and records of the plurality of synthesis containers, wherein the host computer contains links between the data fields whereby the synthesis container is determined by reading data contained within the memory of the matrix with memory.

7. The system of claim 6, wherein the plurality of data fields further includes a data field comprising records containing identifying or historical information pertaining to the matrix with memory.

8. The system of claim 2, further comprising:

a feeding means for separating a single matrix with memory from a plurality of matrices with memories and disposing the single matrix with memory within the reading range of the identification station for identification of the single matrix with memory, wherein the feeding means is connected to the positioning means.

9. The system of claim 8, further comprising a removable drawer disposed adjacent the positioning means for containing an array of synthesis containers, wherein the positioning means has a range of movement adapted to cover the array of synthesis containers in the drawer.

10. The system of claim 8, wherein the feeding means comprises a vibratory feeder.

11. The system of claim 8, further comprising a programmable logic controller for controlling communication between the host computer and each of the identification station, the positioning means, and the feeding means.

12. The system of claim 11, wherein the programmable logic controller is programmed:

(a) to receive a feed signal from the host computer and direct the vibratory feeder to advance the matrix with memory to the identification station, and

(b) following identification of the matrix with memory, to receive a location signal from the host computer and direct the positioning means to place the matrix with memory in the pre-determined synthesis container.

13. The system of claim 11, wherein the positioning means further comprises:

a first motor attached to a first arm for moving along one of an X-axis and a Y-axis; and

a second motor attached to a second arm that is further attached to the first arm for moving along the other of the X-axis and the Y-axis.

14. The system of claim 12, further comprising a feedback means for providing feedback to the programmable logic controller to monitor the identification station, the positioning means, and the feeding means.

15. The system of claim 1, wherein the matrix material is of a size such that at least one dimension is no more than about 20 mm, or the matrix material is in the form of a container or solid surface used for chemical syntheses, or the matrix material comprises a plurality of solid support particles contained within a microvessel.

16. The system of claim 15, wherein the remotely-accessible memory is an electronic memory.

17. The system of claim 15, wherein the remotely-accessible memory is an optical memory.

18. The system of claim 17, wherein the optical memory is an optically-readable symbol imprinted on the matrix material.

19. The system of claim 18, wherein the optically-readable symbol is a bar code.

20. The system of claim 19, wherein the bar code is a two-dimensional bar code.

21. The system of claim 15, wherein at least one surface of the matrix material or portion thereof that is sufficient to serve as a support is derivatized for linking molecules or biological particles.

22. The system claim 15, wherein the matrix with memory further comprises a molecule, a biological particle, a mixture of molecules, a mixture of biological particles, or a mixture of both molecules and biological particles.

23. The system of claim 15, wherein the matrix material comprises a plurality of solid support particles contained within a structure with a cavity formed therein for retaining the plurality of solid support particles and having at least one porous wall enclosing the cavity for permitting the building block to enter the cavity.

24. The system of claim 15, wherein the matrix with memory is about 20 mm or less in size in the largest dimension.

25. The system of claim 15, wherein the matrix material comprises an elongated tube.

26. The system of claim 15, wherein the elongated tube has two ends and walls of porous or semi-permeable non-reactive material and the two ends are sealed.

27. The system of claim 24, wherein each solid support particle is about 50 μ

m-200 μ

m in its largest dimension or smaller.

28. The system of claim 15, wherein the matrix material further comprises associated luminescent moieties.

29. The system of claim 1, wherein

at least a portion of the matrix material is treated to render it suitable for use as a support matrix for molecules or biological particles.

30. The system of claim 1, wherein the remotely-readable memory comprises an optically-readable symbol imprinted on the matrix material, wherein:

the optical memory is readable by an optical detector;

the optical memory is in contact with or is part of the matrix material;

at least a portion of the matrix material is treated to render it suitable for use as a support matrix for molecules or biological particles.

31. The system of claim 1, where a surface or a portion thereof of the matrix material is radiation grafted with a monomer that is derivatized for linking molecules or biological particles.

32. The system of claim 2, wherein the cue includes an audible signal.

33. The system of claim 2, wherein the cue includes a visually-detectable signal.

34. The system of claim 2, wherein the visually-detectable signal is light from a light emitting diode and the diode is selectively illuminated to indicate a proper destination or an improper destination.

35. The system of claim 15, wherein the matrix material is in the form of a cube or other parallelopiped having an outer surface wherein at least a portion of the outer surface has been treated for linkage of molecules or biological particles.

58. The system of claim 1, wherein the molecule or biological particle is selected from the group consisting of monomers, nucleotides, amino acids, small molecules, antigens, antibodies, ligands, proteins, nucleic acids, phage and viral particles, and cells.

36. An automated system for use in solid phase synthesis comprising:

a plurality of matrices with memories, each matrix with memory comprising a combination of a matrix material for supporting solid phase chemical synthesis and a remotely-accessible memory containing encoded data for uniquely identifying each matrix with memory;

a vibratory feeder having a bowl for retaining a plurality of matrices with memories, the vibratory feeder having an output for feeding matrices with memories out of the bowl;

a supply tube having a first end disposed at the output for receiving matrices with memories, and a second end;

a turnstile disposed at the second end of the supply tube adapted to receive the matrices with memories, the turnstile being rotatable to isolate a single matrix with memory for release from a turnstile release;

a positioning tube disposed at the turnstile release for receiving the matrix with memory;

a releasable stop for temporarily retaining the matrix with memory within the positioning tube;

an identification station adjacent the releasable stop having an antenna adapted to interrogate the matrix with memory when the releasable stop is activated;

a plurality of synthesis containers, each synthesis container adapted for receiving one or more selected matrices with memories, each synthesis container containing a solution comprising a building block for solid phase synthesis on the selected matrices with memories;

a positioner adapted for placing the matrix with memory into a pre-determined synthesis container of the plurality of synthesis containers; and

a computer in electrical communication with the identification station, the positioner, the turnstile and the releasable stop, the computer having a database comprising a plurality of synthesis sequences, each synthesis sequence comprising a plurality of steps for synthesizing a molecule or biological particle from a plurality of building blocks, wherein each synthesis sequence corresponds to an identified matrix with memory and the computer identifies the predetermined synthesis container for placement of the identified matrix with memory for performing the next step in the synthesis sequence.

37. The automated sorter of claim 36, wherein the releasable stop comprises a gating solenoid having a plunger adapted to penetrate the positioning tube so as to prevent the passage of the matrix with memory when the solenoid is activated.

38. The automated sorter of claim 37, wherein the positioner further comprises a first linear translator connected to the positioning tube for translating the positioning tube to a location over the designated container.

39. The automated sorter of claim 36, wherein the positioner comprises:

a first arm for providing movement along one of an X-axis and a Y-axis; and

a second arm connected to the first arm for providing movement along the other of the X-axis and the Y-axis, the second arm being attached to the positioning tube;

wherein movement along the first arm and the second arm provide for two dimensional positioning of the positioning tube over the designated container.

40. The automated sorter of claim 39, further comprising:

a programmable logic controller in electrical communication with the positioner;

a first motor driver in electrical communication with the programmable logic controller for controlling translation of the first arm in response to a first signal from the programmable logic controller;

a second motor driver in electrical communication with the programmable logic controller for controlling translation of the second arm in response to a second signal from the programmable logic controller; and

a solenoid for retracting the releasable stop to release the memory system in response to a third signal from the programmable logic controller.

41. An automated system for use in solid phase synthesis comprising:

a plurality of matrices-with-memories, each matrix-with-memory comprising a matrix material for supporting solid phase synthesis in physical contact with a remotely-accessible memory containing encoded data for uniquely identifying each matrix with memory;

an array of synthesis containers adapted for receiving one or more selected matrices with memories, each synthesis container containing a solution comprising a building block for solid phase synthesis on the selected matrices with memories;

a feeder having a vessel for receiving the plurality of matrices-with-memories and a vessel outlet for feeding the plurality of matrices-with-memories one at a time out of the vessel;

a singulator means connected to vessel outlet for receiving the plurality of matrices-with-memories and dispensing a single matrix-with-memory of the plurality of matrices-with-memories;

a positioning means disposed for receiving the matrix-with-memory dispensed from the singulator means, wherein the positioning means holds the matrix-with-memory until a release signal is received;

a gating means within the positioning means responsive to the release signal for retaining the matrix-with-memory within the positioning means and releasing the matrix-with-memory when the release signal is received;

an identification station disposed within a reading distance of the positioning means for reading data stored within the matrix-with-memory when the matrix-with memory is held within the positioning means, wherein the identification station generates a data signal comprising information corresponding to the matrix-with-memory;

a translator for moving the positioning means in response to a positioning signal for positioning the matrix-with-memory over a pre-determined synthesis container within the array of synthesis containers; and

a system controller in electrical communication with the positioning means, the gating means, the translator and the identification station, the system controller having a database comprising a plurality of synthesis sequences, each synthesis sequence comprising a plurality of steps for synthesizing a molecule or biological particle from a plurality of building blocks and each synthesis sequence corresponding to an identified matrix with memory, wherein the system controller receives the data signal and associates the information therein with a corresponding synthesis sequence of the plurality of synthesis sequences to determine a next step within the synthesis sequence, identifies the pre-determined synthesis container corresponding to the next step within the synthesis sequence, generates the positioning signal for moving the positioning means to the pre-determined synthesis container, and generates the release signal for releasing the matrix-with-memory from the positioning means into the pre-determined synthesis container.

42. An automated system for use in solid phase synthesis comprising:

a plurality of matrices with memories, each matrix with memory comprising a combination of a matrix material for supporting solid phase synthesis and a remotely-accessible memory containing encoded data for uniquely identifying each matrix with memory;

a plurality of synthesis containers adapted for receiving one or more selected matrices with memories, each synthesis container containing a solution comprising a building block for solid phase synthesis on the selected matrices with memories;

a singulator for advancing a single matrix with memory;

a dynamic dropper for receiving the matrix with memory from the singulator;

an identification station for reading data stored within the matrix-with-memory when the matrix-with memory is held within the dynamic dropper, wherein the identification station generates a data signal comprising information corresponding to the matrix-with-memory;

a means for positioning the dynamic dropper above a pre-determined synthesis container corresponding to a next step in a solid phase synthesis sequence, wherein the matrix with memory is dropped into the pre-determined synthesis container; and

a computer in electrical communication with the identification station and the positioning means and having a database comprising a plurality of synthesis sequences, each synthesis sequence comprising a plurality of steps for synthesizing a molecule or biological particle from a plurality of building blocks and each synthesis sequence corresponding to an identified matrix with memory, wherein the host computer determines a destination of the identified matrix with memory, the destination comprising a pre-determined synthesis container of the plurality of synthesis containers.

43. An automated system for use in solid phase synthesis comprising:

a plurality of matrices-with-memories, each matrix with memory comprising a combination of a matrix material for supporting solid phase synthesis and a remotely-accessible memory containing encoded data for uniquely identifying each matrix with memory,

a feeder having a vessel for receiving the plurality of matrices-with-memories and a vessel outlet for feeding the plurality of matrices-with-memories one at a time out of the vessel;

a singulator means connected to the vessel outlet for receiving the plurality of matrices-with-memories and dispensing a single matrix-with-memory;

a positioning means disposed for receiving the matrix-with-memory dispensed from the singulator means, wherein the positioning means holds the matrix-with-memory until a release signal is received;

a gating means within the positioning means responsive to the release signal for retaining the matrix-with-memory within the positioning means and releasing the matrix-with-memory when the release signal is received;

an identification station disposed within a reading distance of the positioning means for reading data stored within the matrix-with-memory when the matrix-with memory is held within the positioning means, wherein the identification station generates a data signal comprising information corresponding to the matrix-with-memory;

a plurality of synthesis containers, each synthesis container adapted for receiving one or more selected matrices with memories after release from the positioning means, wherein each synthesis container contains a reagent comprising a building block for solid phase synthesis on the selected matrices with memories;

a controller including a database comprising a plurality of synthesis sequences, each synthesis sequence comprising a plurality of steps for synthesizing a biological particle or molecule; and

an X-Y translator for moving the positioning means along an X-axis and a Y-axis in response to a positioning signal for positioning the matrix-with-memory over a pre-determined synthesis container containing the reagent corresponding to the next step in the synthesis sequence;

wherein the controller receives the data signal and associates the information therein with the next step in the solid phase synthesis sequence, for identifying a location of the synthesis container corresponding to the next step within the synthesis sequence, for generating the positioning signal for moving the positioning means to the synthesis container, and for generating the release signal for releasing the matrix-with-memory from the positioning means into the synthesis container.

44. The automated system of claim 43, wherein the vessel outlet includes an orientation means for feeding the plurality of matrices-with-memories to the singulator means with a pre-determined orientation.

45. The automated system of claim 43, wherein the singulator means comprises:

a body having an angled bore disposed therethrough for receiving the matrix-with-memory so that the matrix-with-memory slides from an upper end to a lower end of the angled bore; and

a gating solenoid in electrical communication with the controller, the gating solenoid for gating the release of the matrix-with-memory from the angled bore in response to a gating signal generated by the controller.

46. The automated system of claim 45, wherein the gating solenoid comprises two gating solenoids, each gating solenoid receiving a separate gating signal from the controller.

47. The automated system of claim 45, wherein the singulator means further includes a plurality of position sensors for sensing a position of the matrix-with-memory within the angled bore and for generating and providing a plurality of position signals to the controller for indicating the position within the angled bore.

48. The automated system of claim 47, wherein the controller receives a first position signal of the plurality of position signals and generates the gating signal in response thereto.

49. The automated system of claim 47, wherein the identification station is disposed near an exit of the singulator means, and the controller generates a command for activating the identification station in response to a second position signal of the plurality of position signals.

50. The automated system of claim 43, wherein the singulator means comprises:

a turnstile having at least one slot for receiving the matrix-with-memory, the turnstile being rotatable around an axis so that the at least one slot for receiving the matrix-with-memory can be rotated to an exit position for releasing the matrix-with-memory; and

a drive means in electrical communication with the controller for rotating the turnstile in response to a rotation signal generated by the controller.

51. The automated system of claim 43, wherein the controller comprises a programmable logic controller and a host computer for running a plurality of software programs including a synthesis manager program for associating the information for the matrix-with-memory with one or more steps in a corresponding synthesis sequence.

52. The automated system of claim 51, wherein the plurality of software programs includes a sorter interface program for controlling the programmable logic controller in coordination with the synthesis manager program.

53. The automated system of claim 51, wherein the synthesis manager program generates a searchable record corresponding to each matrix-with-memory and each compound to be synthesized in the corresponding synthesis sequence.

54. The automated system of claim 53, wherein the host computer further includes a graphic display, and the synthesis manager program generates a display of a map of the plurality of matrices-with-memories and the container associated with each matrix-with-memory.

55. The automated system of claim 53, wherein the host computer further includes a graphic display, and the synthesis manager program generates a display of the building block for each compound to be synthesized in the corresponding synthesis sequence.

56. The automated system of claim 52, wherein the plurality of software programs includes a template program for identifying a pre-determined layout for the plurality of synthesis containers, the template program being interactive with the synthesis manager program and the sorter interface program.

57. The automated system of claim 52, wherein the plurality of software programs includes a calibration program for calibrating movement of the X-Y translator for a plurality of increments along the X-axis and the Y-axis.