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Umted States Patent [191 [11] Patent Number: Mezei, Louis M. et a1. ' [45] Date of Patent: [54] USER CONTROLLED OFF-CENTER LIGHT 4,253,846 4,255,788 3/1981 Schwartz et al. 364/413.07 . . . .. LING AND REACI'ION [75] Inventors: Mezei, Louis M_, Fremont; Bradley ‘ 4,268,268 5/1981 Blum 4,319,882 3/ 1982 Sharma ........... .. . .... .. .. 4,451,433 5/1984 Yamashita et a1. 422/63 4,478,094 10/1984 Solomaa et al. .. 73/ 863.32 4,556,641 12/1985 Kano et al. Stephen J. Moehle’ both of Berkeley; 4,580,895 4/1986 Patel . . . . . . . . . . . . . . 436/52 436/63 422/73 x . . . .. 356/39 Brent S. Noorda, Pleasant H111, . 4, 665 , 553 5 /l987 4,608,246 8/1986 Bayer et a1. Gershman et al. .. 424/11 356/39 X J°sePh T- W‘d‘mas’ Berkeley; James A- Zei?in, P‘edm‘mt’ all 01' Cahf- 4 678 894 4,683,120 7/1987 Shafer ............. .. 7/1987 Meserol et a1. 364/416 x ....... .. 422/72 . ' ' ' 4,713,348 Asslgnee. Cetus Corporation, Emeryvllle, Cahf. Appl. No.: 906,101 [22] Filed: Smythe et a1. ...................... .. 436/53 s_ Albom, Richmond; coppock, Stan; - , [73] [21] Oct. 10, 1989 ABSORBANCE READING ADJUSTER IN A AND 3/1981 4,873,633 . 12/1987 Ullman ........ .. 436/501 4,719,087 V1988 Hanaway .... n 422/102 4,727,033 2/1988 Hijikata et a1. ..................... .. 356/39 Sep. 11, 1986 Primary Examiner-Clark A. Jablon Attorney, Agent, or Firm-Ronald C. Fish; Kevin R. Kaster; Albert P. Halluin [57] ABSTRACT A host computer controls a plate reader which optically Related US. Application Data [63] Continuation-impart of Ser. No. 788,998, Oct. 18, 1985, abandoned’ [51] Int. Cl.4 ................... .. G01N 33/48; GOlN 21/01; reads the results between donor samples and reagents. G01N 33/80; G01N 35/02 The system provides the user with the ability to change [52] US. Cl. ............................... .. 364/413.08; 356/39; 356/442; 422/73 [58] Field of Search ---------------- -- 364/416, 555; 422/73; 356/39, 4421 440 [56] References Cited the position of off-center light absorbance readings to maximize the power of the machine to discriminate between positive and negative agglutination reactions. In addition, the user can tailor thresholds for absorb ances used to distinguish between positive, negative and “no type determined” reactions. U.S. PATENT DOCUMENTS 4,130,395 12/1978 Chryssanthou ..................... .. 424/11 7 Claims, 4 Drawing Sheets W HAVl C ABSORB ANC E POSITIVE REACTIONS 1,511 ‘POSITIVE ACTUAL ABSORBANCE AT POSITIVE THRESHOLD .1 1 1,. NEGATNE NEGATlVE THRESHOLD ‘maesuow REACTIONS Atrgls’gll-? ACTUAL ABSORBANCE AT NEGM’NETt-RESHOlD ABSORB ANCE POSITIVE REACTION AGGLUTINA TION NUMER OF NTD'S AT CURRENT THRESHOLD SETTING WELL C‘ENTER ""11 \ meant?“ WALL ‘~—' U DEFINA READINBéE POSITIONS 131141958811“ WELL WALL US. Patent HOST 0ct.10,1989 Sheet 1 0f 14 [/20 4,873,633 MAINFRAME CPU DEC PDP II 30 PRINTER / (32 (22 KEYBOARD 28 CENTRIFUGE ( PLATE READER TERMINAL 24J ORBITAL /34 SHAKER I LIQUID HANDLER 26 LIQUID HANDLER AND PLATE #2 HANDLING MECHANISM FIG. I LIQUID HANDLER . . . #N US. Patent Oct.10,1989 Sheet 2 of 14 4,873,633 US. Patent Oct. 10,1989 I Sheet 3 0114 4,873,633 LOGON FOR SYSTEM STARTUP USER _/lO2 PRIV TABLE FIVE MAIN MENU #104 LOG ON FOR SYSTEM sTARTUP REAGENT DATA HANDLING "54 ('40 QUALITY CONTROL PROCEDURES LIQUID HANDLING PARAMETER A00 DEFINITION PLATE READING EDIT RESULTS _/-|7O (I90 REsULTs SUMMARY READER CALIBRATION /— O0 5 ( 300 SYSTEM CONFIGURATION FILE OPERATIONS "400 . ('38 IM MUNOASSAY F IG . 3 US. Patent Oct. 10, 1989 Sheet 4 0f 14 4,873,633 LIQUID HANDLING MENU A06 (H2 ' BROM PROCEDURE FOR 3 ABO/Rh TESTING: '08 [H6 PROCEDURE FOR ABO/Rh TESTING= SAMPLE HANDLING REAGENT HANDLING QROCM PROCEDURE FOR no, ABO/Rh TESTING ,-II4 L SELECT CRITERIA "'8 FOR REPEAT LIQ. HANDLING I (120 (I30 OPTIONS F OR SPECIFYING WBN'S NTD S OF SAMPLES FOR I REPEAT LIQ. HANDLING [T122 ‘ DISPLAY OF NTD'S —‘ PROCESSED BY WBN _ I PRINT (I32 PROC. FOR REPEAT LIO. HANDLING Q__l FOR SAMPLES PROC. FOR REPEAT A24 LIQ. HANDLING _ ('34 FOR NTD S I B BROM PROC. FOR REPEAT YES LIO. HANDLING ( 126 PROC. FOR REPEAT YFEOQA LIO. HANDLING OF NTD'S= SAMPLE (I28 _ SAMEEZAN‘éE'PINE HANDLING FOR SAMPLES: SAMPLE HANDLING ('36 SAME: REAGENT HANDLING i FIG. 4 US. Patent Oct. 10, 1989 Sheet 5 of 14 4,873,633 QUALITY CONTROL MENU NO BROMELIN PROCEDURE FOR202 PRETREAT CHECKING REAGENT CELL SUSRENSIONS (I64 (204 ABSORB OF REAGENT BROMFEEQ QEIXEEIEQRCCETECSISSNS CELL SUSP FRET A SAMPLE HANDLING ‘ TABLE a MENU RESULTS NOT ACCEPU[ ] VIEW I N67 ACCEPTED (#68PROC. FOR CHECKING (206 QC ADJUST CELL PRINT CHART » SUSPENSION REAGENT CELL SUSF; J REAGENT HANDLING L DO QC LIQ. | HANDLING BI PLATE READING . QC OPERATOR OK ID LOGON NO LIO. HANDLING ID OF QC BROM SAMPLES I18; _I LI42 LI44 (‘I46 SIEOCEDURE FOR — BROM FROG FOR QC; YES SAMPLE HANDLING RESULTS OF Qc PROcEDURE /-'52 I48/ I-IEJ PROC. FOR QCI REAGENT HANDLING PERFORM QC LIQ. HANDLING 150/ "'53 L a cOLLEcT ~ ABS. DATA NO BROM PROG. FOR H69 CHECKING DONOR CELL SUSP 1 fl?! PROc. FOR BRQM CHECKING I75\ DO QC LIQ HANDLING 'a fl73 PROc. ‘FOR CHECKING DONOR GELL DONOR CELL E‘IISNPISLSIRIIPLE RIEESEEN‘TSIS’IIN‘DLING I PLATE READING CHECKING REAGENT IGIJ PROc. FOR SAMPLE EAIIEIINGFOR IrERGO To EICHEER REAGENT TITER _—____I 58 FIGSB HANDLING \IGT ~—l63 FIG. 5A US. Patent To FIGSAT 0ct.10,1989 Sheet 6 of 14 4,873,633 FROM ‘FIG. 5A QUALITY CoNTRoL CoNT' D 1 VIEW TITER GRAPHS '62 I (I66 (‘I70 ABSORBANCE OF DoNoR CELL sUsPENsIoNs ‘ ‘ RESULTS NOT ADJUSTING DONOR CELI ‘ ACCEPTED SUSP ACCERTI [PRINT )—1 CHART VIEW (QC/I75 77 I REAGENT DATA HANDLING I56w CURRENT REAGENT DATA TABLE REAGENT DATA FILE l— PRINT J ('58 ACCESS To REAGENT DATA MODULE LOGON UsER RRIvILEGE TABLE FIVE LACCESS NoT OK ACCESS OK (I60 CURRENT REAGENT DATA TABLE REAGENT DATA FILE [MAIN MENU‘ F166 EB LPRINTJ US. Patent 0a. 10, 1989 Sheet 7 0f 14 4,873,633 PLATE READING MAIN MENU EDIT RESULTS REREAD ROW fws AUTOMATICALLY I LOGON TO ENTER OR EDIT ABO/Rh TEST RESULTS MANUALLY NO ACCESS |_ ACCESS OK ENTER 0R EDIT RESULTS MANUALLY ' ENTER RESULTS MANUALLY FOR WBN I EDIT RESULTS FOR WBN LOGON TO ENTER OR EDIT DU TEST RESULTS MANUALLY /I72 ACCESS OK ENTER OR r-I74 EDIT DU RESULTS FROM CURRENT LIST No ACCESS I= LOGON TO ENTER OR EDIT MISC. TEST RESULTS M A NUALLY _ NOACCESS L I SELECT ACCESS TEST TO OK ENTER EDIT RESULTS I I ENTER OR EDIT RESULTS FOR SELECTED TEST l ENTER REMARKS MAIN MENU FIG. 7 US. Patent Oct. 10,1989 4,873,633 Sheet 8 of 14 RESULTS SUMMARY (2H WBN LIST FOR SPECIFIED DISPLAY OF WBN “20' T" GROUP AND TEST RESULTS TYPE I (2I3 DISPLAY OF T’ TEST STATUS 207 @ (215 DIRECTORY OF , " PLATE INFO TABLE EEEIJ DISPLAY OF UNVERIFIED Rh NEGATIVE LIST E FOR PLATE 4? _ I j DIRECTORY OF ANTI BODY SCREEN PLATES POSITIONS FOR EZIT l PLATE =I=I= E DISPLAY OF ABO/ Rh TOTALS DISPLAY OF DATA J __|93 ~223 E (225 LIST OF OMITTED CAROUSEL F POSITIONS _.IPRINT LAB TEST RECORD E FIG. 8 US. Patent 4,873,633 ‘Sheet 9 0f 14 Oct. 10, 1989 FIG. 9A 502 PLATE READER NO CALIBRATION LOGON‘ Acct-35 READER CALIBRATION _,_ 504 MENU @ ( 506 —-| DETERMINING THRESHOLD VALUES I (“508 BROM PROC. FOR LIQ. HANDLING OF THRESH. DETERM PLATES NO r510 BROM PROC, FOR SAMP HANDLING YES FOR THRESH. DETERM. PLATES PROC. FOR REAGENT HANDLIMS . OF THRESHOLD DETERM PLATES I SPECIFY HISTOGRAM TO REVIEW _ . I EVAL. TEMP THRESHOLD VALUES MENU MANUAL ENTRY OF SAMPLE INFO FOR EVAL OF TEMP THRESH. VALUES L DISPLAY OF NTDS DERIVED FROM THRESH. DETERM. PROCESS I NOT OK DIS P LAY OF DISCRE PANCIES BETWEEN ENTERED SAMPLE VALUES AND THRESHOLD 2K DETERM. PROCESS 520i 522A VERIFY UPDATE OF F— THRESHOLD NO TO r98. FROM FIG. 9B VALUES REPLACE I I REPLACE REVIS E H I STOGRAMS 524 532 L___________I OK INoT OK (528 SPECIFY HISTO GRAMS TO REVIEW I ( 5530 REVIEW OF EDITED THRESHOLD VALUES MENU | {2 3 4 5 US. Patent TO FIG. 9A Oct. 10,1989 Sheet 10 0f 14 4,873,633 FROM FIG. 9A I {540 BROM PROCEDURE FOR PLATE READING YES POSITION DETERMINATION f544 SAME- REAGENT HANDLING (542 RRoc. FOR PLATE BROM READING POSITION NO DETERMINATION: LIQ. HANDLING V (548 STEPS OFF CENTER DETERMINATION L546 (554 VERIFY UPDATE OF PLATE READING POSITION FIG. 9B US. Patent L06 0 N 0a. 10, 1989 NO ACCESS Sheet 11 0f 14 4,873,633 MAIN MENU FILE OPERATIONS MENU TRANSFER COMPLETED r404 RESULTS TO DISK 408 _ REVISE STORED DATA I RESET DATA FOR SPECIFIED WBN ENTRY ‘DELETE DATA lI RESET DATA FOR SPECIFIED PLATE ENTRY 4IO IL——I DELETE ENTRIES DELETE ENTRIES DELETE BY DATE BY PLATE BY WBN K I k 4II BACKUP FILES 4I2 406 DISPLAY DATA STATISTICS FORMAT FLOPPY DISK/—-402 MAIN MENU FIG. IO ( 4I3 DELETE IF TRANSFERRED TO MAINFRAME IL4I4 US. Patent Oct. 10, 1989 Sheet 12 Of 14 4,873,633 NO ILO GON F OR SYST EM c ONFIGURATIONI'——>@ ACCESS ACCESS OK SYSTEM CONFIG. MENU (304 (306 MODIFY CURRENT TEST FORMAT MODIFY ADO/Rh LIQ. HANDLING PROC. I I @ (‘302 CURRENT TABLE OF USER ID/ SYSTEM SECURITY INFO I (30s —-IMODI FY MAINFRAME HANDSHAKE] 3270 EMULATION SNA EMULATION SPECIFY HANDSHAKE FOR RS232 PROTOCOL I I I —-IsEI_EcT LAB REPORT FORM |~3IO J MODIFY IMMUNOASSAY FORMAT _,3|4 AN D LIO. HANDLING I _-IMODIFY ERROR HANDLING METHODI~3I2 I —-IMODI FY HARDWARE OOMPLEMENT] I FIG. II US. Patent Oct. 10, 1989 Sheet 13 0f 14 4,873,633 0/0 OF TOTAL wELLS HAVING EAcH ABSORB ANCE I I III II PPM“ II WES‘? POSITIVE 5'9 yPOSITIVE REACTIONS xxxx’TfL23 I II ¥_“’—/ NEGATIVE NEGATIVE THRESHOLD THRESHOLD REACTIONS YYYY/ 525 ACTUAL ABSORBANCE AT POSITIVE THRESHOLD ASAIVIRLE W POSSIBLE 22 2’/5.2 I ACTUAL ABSORBANCE AT NEGATIVE THRESHOLD NUMBER OF NTD'S AT CURRENT THRESHOLD SETTING WELL ABSORBANCE CENTER POSITIVE REACTION AGGLUTINA TION I POS TION ABSORBANCE READING POSITIONS FIG. ISA WALL READING I RELATIVE TO H4 WELL WALL USER DEFINABLE READING POSITIONS FIG. I38 US. Patent Oct.10,1989 Sheet 14 0f 14 4,873,633 ABSORBANCE I<——|65 I | l | I I : 2 : 4 : 8 | ' : i I6 32 DTLuTToN OR TITER ABSORBANCE MAX. , . _ - ' ' " MEAN MIN I23 WBN I23 135 620 3o---- eo--- 90 DAY KNOWN EXPERIMENTALLY WBN TYPE DETERMINED BLOOD TYPE 23 TEMPLATE FAPOS BPOS o BPOS++———++ APos +——+++—+' o FIG. l6 — — + — — — + + 1 4,873,633 2 not subject to automated liquid handling. Test results USER CONTROLLED OFF-CENTER LIGHT ABSORBANCE READING ADJUSTER IN A LIQUID HANDLING AND REACTION SYSTEM This application is a continuation-in-part of Ser. No. 788,998, ?led Oct. 18, 1985, now abandoned. CROSS REFERENCE TO MICROFICHE APPENDIX There is included herewith micro?che appendices, consisting of 10 micro?che and 461 frames, including the hex format object code controlling the various pro from these antibody screen tests and other tests done by hand must be recorded for each donor’s blood for which these tests are done. Further, testing procedures for blood typing can change over time in that different dilution values are often needed for different batches of reagents to get the proper absorbance readings. Fur~ ther, the amount of dilution of the red blood cells should be optimized. The data for the optimal dilution values must be recorded. Further, other process parameters of the blood typing sequence should be subject to customization to enable the various users to customize their testing procedures to conform to local practice. This data de?ning the cessors in the system. 15 characteristics of each step, such as the amount of each BACKGROUND OF THE INVENTION sample to be placed in each well, which wells in which to place samples, the amount of dilution in certain steps, The invention pertains to the ?eld of systems for the wells to place reagents in, the number of mixes at automatically performing blood typing operations, and, various points in the process and various other criteria, more particularly, to systems for controlling automated liquid handling apparatus to do blood typing and for should be subject to customization, should be easily interpreting the results from a plate reader and for man changeable, and should be remembered by some mecha aging the data generated by the liquid handling appara nism so that it can be automatically invoked each time a tus and the plate reader apparatus and for printing vari procedure is performed without having to look it up every time. Of particular importance is the amount of dilution of each particular reagent which should be used ous reports. Modern blood banks must perform thousands of 25 blood typing and antibody screening operations and manage the data resulting from such tests. These opera to optimize the absorbance for that reagent and to con serve the amount of reagent used. These reagents are tions involve the handling of thousands of samples of donor blood, the pipetting of reagents into samples of often quite expensive. the plasma and red blood cells from the donor blood accountability system such the quality of data generated in the testing is consistently high and the persons per forming the testing can be determined. For example, it and the optical reading of the wells containing the donor samples and the reagents to determining the pat Further, it is useful to have a quality control and is useful to know the expiration data for all the reagents in stock and the associated absorbance values for each constitutes a template which characterize the blood as 35 reagent lot over the period of its usage. It is also useful to compare the test results for known sample types to being from a particular blood group and having a par the results that should have been obtained as a check on ticular Rh factor. Each test of each donor’s blood in the accuracy of the system. It is also useful to be able to volves pipetting of multiple samples of that donor’s generate reports on daily or monthly activities to deter plasmas into multiple wells and pipetting of multiple mine the amount of certain types of blood in stock and samples of diluted red blood cell samples into multiple where it can be found. The number of “no type deter wells. Multiple diluted reagents are then added to the multiple wells containing plasma and red blood samples mined” test outcomes (hereafter NTD). Thus, a large amount of data in the form of process and various reactions either occur or do not occur de controlling parameters and test results are involved in pending upon the blood type and the reagent in each well. Typically, these reactions manifest themselves as 45 blood typing and antibody screening operations, and a large amount of record keeping for this data is involved. clumps of protein in the bottom of the well for a posi Therefore a need has arisen for a system which can tive reaction and no clumps for negative reactions, the tern of positive and negative responses to various rea gents. The pattern of positive and negative responses pattern of positive and negative reactions determines perform these thousands of liquid handling steps reli the blood type. The blood type can be determined by placing the multiple wells under a strong light source and reading ably and tirelessly and which can handle the thousands the optical absorbence, i.e., the amount of light which gets through the bottom center of the well. Typically absorbance readings are taken on both sides of the cen of data records which characterize the liquid handling and which constitute the data record for each donor. Further, such a system should be access controlled and implement accountability. It should also ease quality control operations and maintain data useful in quality ter, bottom of each well, and the results are compared 55 control efforts. Such a machine should also maintain all test results and be able to communicate them to a main to certain threshold criteria for absorbance. The com frame computer for permanent storage or allow them to parison of the readings will indicate the presence of a be archived onto permanent magnetic storage. Such a clump at the bottom of the well (low absorbance in the machine should also enable the automated generation of off center readings) and therefore a positive reaction or the absence of a clump and therefore a negative reaction 60 reports for management. (high absorbance on both off center readings). SUMMARY OF THE INVENTION Clearly, the process of blood typing of thousands of The invention is a system for performing a host of donor samples involves many thousands of liquid .han functions which aid in the operational work and man dling steps and the generation of many thousands of absorbance readings for the multiple wells devoted to 65 agement of a blood bank. The invention consists of various off the shelf components and a custom designed each donor. Further liquid handling steps are involved liquid handler combined in one system, all controlled by in automated antibody screening, and some special tests may have to be done by hand on the blood which are a comprehensive software resident in a host computer 3 4,873,633 4 which allows the user to control blood typing and other host is also coupled to an orbital shaker and a centrifuge liquid handling operations in the liquid handler and to for re-suspension and re-separation of liquids and sam ple cells during some system operations. manage the data which results therefrom. The system automates: much of the liquid handling involved in The host software also contains routines for imple processing donor samples for ABO/Rh blood typing tests and antibody screening; sample and plate identi? menting quality control operations. Such operations include verifying test procedures by processing a small number of known blood type samples and checking the experimental data against the known results, and verify ing that the dilution of cell suspension reagents and donor cells produce absorbances in the proper range. The system can also test the titer of antibody reagents. The data management functions of the invention cation using a bar code reader; plate reading for the ABO/Rh tests; record keeping for quality control data and test results for individual samples; access to infor mation on test status; and printing of laboratory records and transmission of information to a mainframe com puter for on-line storage. The physical system includes a Digital Equipment Corporation PDP-ll MICRO with a 10 megabyte hard include the management of data in the form of: test procedure parameters for liquid handling, plate reading disk and a VT220 terminal. The host system 20 runs a commercially available operating system. The control software is resident in the host. It is coupled to a custom designed liquid handling system which has several mi croprocessors resident therein which control various stepper motors, bar code read heads and monitor vari 20 ous sensors. The process controlling parameters to con and histograms; quality control data consisting of rea gent lot numbers and expiration dates with associated absorbances of these reagent lots over the period of their usage; test results including blood groups and probable type; manually added data from such tests as DU tests, antibody screens, hepatitis, HTLV III, CMV, RPR and other tests; and test status for the day of a trol operations of the liquid handler come from the host range of parameters such as the number of A+ samples computer after being de?ned by the user at the host that have been processed, the number of NTD’s during terminal. Once the process parameters are loaded, the resident software in the liquid handler controls various 25 the day and the status of testing for particular plates. transfers of donor blood plasma and red blood cells to BRIEF DESCRIPTION OF THE DRAWINGS various wells in plates loaded by an automated plate FIG. 1 is a block diagram of the interconnection of reader. The resident software also controls dilution of the various components of the system. the red blood cells before depositing them in wells and FIG. 2 is perspective view of the liquid handler of the the transfer of speci?ed quantities of various types of system. reagents into the wells containing the donor’s blood FIG. 3 is a flow diagram of the user options for vec plasma and diluted red blood cells. Each plate can hold toring processing to various sub-functions that the sys samples from eight donors, and the donor samples are tem can perform. stored in test tubes in a circular, rotating lazy-susan-like FIG. 4 is a flow diagram for steps performed in al device which is controlled by the software of the liquid 35 lowing the user to customize the process parameters handler. The plates so ?lled are then unloaded by a plate han controlling liquid handling operations. dler device, and manually transferred to a plate reader FIGS. 5A and 5B are a ?ow diagram of the process of Model Autoreader EL309 manufactured by Biotech allowing the user to monitor and control various quality Instruments which is also coupled to the host. The plate 40 control functions. reader then obtains absorbance readings on the wells in FIG. 6 is flow diagram for the process of allowing the the plates after reading the bar codes on each plate. user to create and access various reagent data for use in These absorbance readings for each well on each plate insuring proper processing and improving reliability of are sent to the host with the bar code of the plate for test results. 45 interpretation. FIG. 7 is a flow diagram of the plate reading function The host knows which wells of each plate contain of the system and a flow diagram of the process fol samples from which donors having obtained this infor lowed in allowing the user to edit or enter various re mation by downloading the bar code data read by the sults from testing. liquid handler from the donor test tubes and the plates FIG. 8 is a ?ow diagram for the process followed in that received samples from each tube. The absorbance allowing the user to specify results summaries the user values are interpreted by plotting histograms for the number of samples having each absorbance for each would like to see. FIGS. 9A and 9B are a ?ow diagram of the process that the system follows in allowing the user to calibrate different type of reagent. The user is then allowed to de?ne the threshold values between the positive and negative reactions for each reagent by viewing the histogram and graphically moving an arrow to the de sired threshold value. A “no man’s land” region where the absorbance does not clearly indicate either a posi tive or a negative value is also de?ned, and samples having absorbance values falling in this region are tagged as NTD. The user de?ned thresholds for each type of reagent de?ne “templates” or patterns of posi 55 the plate reader and to adjust the thresholds used by the host computer in interpreting test results. FIG. 10 is a ?ow diagram of the process used by the system in performing various ?le operations. FIG. 11 is a flow diagram of the process used by the system in allowing the user to con?gure the system for a speci?c installation. FIG. 12 is an illustrative histogram for a single rea gent illustrating the meaning of adjusting the thresholds tive and negative reactions which will de?ne each par used in interpreting test results. ticular blood type. The absorbance values for each FIG. 13A shows a typical well bottom with a positive sample are then compared to these templates and typed 65 reaction button. as to their blood group type. FIG. 13B shows a typical graph resulting from a The host is also coupled to a printer, and the user can request various pre-formatted management reports. The positive reaction button. _ 5 4,873,633 6 much of the liquid-handling involved in processing donor samples for ABO/Rh tests and antibody DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT Referring to FIG. 1 there is shown a block diagram of screening, sample and plate identi?cation using a bar code reader, the system of the invention. The system is comprised of 5 plate reading for ABO/Rh tests, record keeping for quality control and test results for a Digital Equipment Corporation PDP-ll MICRO host individual samples, computer 20 which coordinates the activities of the system under the control of the resident software in cluded herewith as the appendices and the user (not shown) who enters commands through a keyboard 22 in response to information displayed by the software on a access to information to test status, and printing of laboratory records and transmission of infor mation to a mainframe computer for on-line storage. ProGroup Automated Blood Typing System Bene?ts terminal 24. The host computer 20 runs the commer The ProGroup TM automated blood typing system is a ?exible system, designed to adapt to the way work is cially available Micro RSX operating system for the PDP 11/23, ll/53 or ll/73 computer versions available from Digital Equipment Corporation. The application already organized in the user’s laboratory. The user de?nes the way he or she wants particular procedures carried out, and ProGroup TM automated blood typing software which implements the functions described herein is included herewith an appendix. The host com system executes these tasks at command. puter is coupled by RS232 serial ports and connecting Automating liquid-handling makes test results more cables to a number of liquid handling and plate handling mechanisms of which liquid handler 26 is an example. 20 reliable, and allows employment of lab workers having less skill. Features like computerized, data management The preferred embodiment of the system can control up techniques and the built-in bar code reader reduce the to 8 liquid handlers. The details of the liquid handlers incidence of operator error. and plate handling mechanism are given in a copending ProGroup TM automated blood typing system does U.S. pat. application assigned to the assignee hereof not require any previous experience with automated liquid-handling equipment, or with computers. entitled “Automated Liquid Handling Apparatus And Process With Plate Handler”, ?led Oct. 18, 1985, Ser. No. 789,945, now abandoned, and another U.S. pat. ProGroup System Overview application entitled “Liquid Manipulation Device and Method”, ?led Jul. 5, 1985, Ser. No. 752,449, now aban 30 doned, both of which are hereby incorporated by refer ence. The object code for the liquid handler disclosed in the above identi?ed patent application has been changed somewhat since the filing date thereof. The latest object code is included herewith in the appendices hereto. The object code appendices are labelled for the particular microprocessor in the liquid handler for There follows more detail on the individual compo nents of the system. Liquid-Handler 5 The liquid-handling portion of the ProGroup TM automated blood typing system consists of the follow ing items. ProGroup TM automated blood typing system liquid which each is intended. The host 20 is also coupled by an RS232'link to a handler 26, shown in more detail in FIG. 2, automates and an orbital shaker 34 are also part of the system but are not connected to the host 20. The centrifuge and four permanently mounted liquid delivery heads (X, Y orbital shaker are commercially available off the shelf components and are used to process the plates after the samples and reagents have been put in the wells by the liquid handler 26 and before the plates are loaded in the plate reader 28. Collectively, the above described sys head 44. One or several of these can be used in the course of a single procedure, since there is no require— ment to remove one head to install another. The X, Y and 63 of several sizes to minimize cross-contamination. tem may hereafter be referred to as the ProGroup TM The ProGroup TM automated blood typing system automated blood typing system or the system. The ProGroup TM automated blood typing system automates many aspects of the blood typing and group 111 and a maximum of 200 ul. the pipetting of donor plasma directly from a carousel 20 of tubes 22 into designated wells of a microplate. The plate reader 28 which optically reads the absorbance ProGroup TM automated blood typing system mixes values for samples in various wells in plates ?lled with donor red blood cells with diluent in designated propor samples and reagents loaded from the liquid handler. tions before transferring the cells to the wells of the The plate reader is commercially available from BioTek microtitre plates 56 and 58. ProGroup TM automated Instruments, Inc. under the designation Autoreader EL309. 45 blood typing system ?lls the ABO/Rh plate 56 and the antibody screening plate 58 with the appropriate rea The host 20 is also coupled by an RS232 link to a gents from a reagent trough 62. The ProGroup TM printer 30 upon which the host 20 prints the various automated blood typing system liquid handler includes reports of which the system is capable. A centrifuge 32 head 28, 12 channel head 42, ?ll manifold 46, and wash head and 12 channel head use disposable pipette tips 24 accurately transfers volumes between a minimum of 10 Two identical bar code readers (not shown) are built ing procedures performed in blood banks. This section 60 into the liquid-handler: one for reading the bar code 21 on the sample tube (actually located above the top sur face of the carousel 20, and the other for reading the bar code 57 and 59 on the microplates (actually located on ProGroup TM Automated Blood Typing System the other sides of the plates). The bar code on the sam Functions 65 ple tube is the basis for the Whole Blood Number (WBN) that identi?es the donor sample. The bar code The major ProGroup TM. automated blood typing on the microplate allows the system to pair the test system functions are broadly de?ned below. ProGroup results from the plate reader with the WBN of the sam TM automated blood typing system automates: provides a brief introduction to the functioning of the system. 7 4,873,633 8 ple donor. The bar code readers can recognize Code the computer, so the user can quickly generate hard Bar or Code 39 labels. copies of laboratory records. Plate Stacker (not shown) holds six 96-well micro plates for ABO/Rh testing. Each plate can accommo date samples from eight donors. These plates are fed into the liquid-handler 26 one by one for processing. Once ?lled, a plate is automatically transferred off the bed 74 of the liquid-handler, into the other rack of the The ProGroup TM automated blood typing system is used by laboratory personnel at three levels: plate stacker. The Technician, who operates the machine on a daily basis, running test and printing results. 7 The Supervisor, who performs quality control proce dures, adds to or revises test data as necessary, handles the storage of information on the ProGroup TM auto ‘ mated blood typing system host computer 20, and data Plate Reader The Plate Reader 28 reads ABO Rh plates automati cally and transmits data points to the ProGroup TM transfers data to a mainframe computer. The Director, who creates the initial framework for the ProGroup. This includes tasks such as defining who will be using the system, how the tests will be carried out, and how test results should be interpreted. The automated blood typing system host computer 20 for interpretation. The results are reported by group and type, or as NTD (No Type Determined) readings. An NTD sample can be reread automatically with the plate Director modi?es this framework as necessary, to re ?ect changes in system users, test protocol, etc.. reader, in which case the results stored on the computer Security Precautions are updated automatically. Alternatively, the row of wells can be examined visually, or the liquid handling 20 The system utilizes a user privilege table and user identi?cation at logon time to guarantee accountability and the reading of that sample can be repeated manu and to insure that only users with the proper authoriza ally. In the latter case, the user inputs information tion have access to certain portions of the program and through the computer keyboard to modify the results of certain data files. the sample processing procedure. The following publicly available documents are in 25 corporated by reference for support material. Orbital Shaker MICRO PDP-ll System Owner’s Manual Digital The ProGroup TM automated blood typing system Equipment Corporation. includes an orbital shaker 34 as part of the system. The Pro/Pette Liquid Handling System User’s Manual Cetus Corporation. shaker is used to resuspend the cells after the liquid handling is complete, and then again after the plate has been centrifuged. The orbital shaker can accommodate up to eight plates. Automated Microplate Reader Model EL309 Opera tor’s Manual, Bio-Tek Instruments, Inc.. Installing and Using the LASO Printer, Digital Equip ment Corporation. Centrifuge The centrifuge 32 can be an instrument such as the Beckman TJ-6R Tabletop Centrifuge. It is used in a known manner on the plates from the liquid handler 26 35 LASO Programmer’s Reference Manual, Digital Equipment Corporation. Orbital Shaker User Manual. ABO/Rh Testing before they are placed in the liquid handler. Host Computer ProGroup TM automated blood typing system auto mates the typing and grouping of donor samples. This The host of the ProGroup TM automated blood typing system is the computer 20, called the host com puter 20. The host computer 20 directs the operation of the liquid handler 26 and the plate reader 28, and man section describes the liquid-handling, resuspension, and plate reading tasks the ProGroup TM automated blood ages the data generated by the other equipment. The user primarily interacts with the ProGroup TM auto mated blood typing system through the host computer 20. The user uses the host computer 20 to select the function the user wants the ProGroup TM automated blood typing system to perform, and to view stored data in tabular or graphic form. The host computer 20 allows the user to modify data that has been created through automatic procedures. The user can also enter addi tional information pertaining to donor samples, such as antibody screen results, or Hepatitis and HTLV III results. The host computer 20 is a DEC MICRO PDP-ll computer with a 10 Mbyte hard disk and a VT 220 typing system performs for ABO/Rh testing. Liquid-Handling Functions The ProGroup TM automated blood typing system liquid handler 26 transfers donor plasma out of the tubes 22 into designated wells of a microplate 56. It also trans fers donor red blood cells into a special predilution block 60, mixes the cells with a de?ned quantity of diluent, then pipettes them into the appropriate wells of the microplate 56. Then the ProGroup TM automated blood typing system fills the plate with the reagents from the reagent plate 62 for the assay. Options Referring to FIG. 3 there is shown the main menu and logon process. Referring to FIG. 4 there is shown the liquid handling process flow chart for the organiza tion of the software if the liquid handling parameter terminal with graphics capability. The computer 20 60 definition menu selection 100 is selected in FIG. 3. FIG. runs the software included herewith as Appendix A on the RSXll operating system which is commercially available from Digital Equipment Corporation. Printer The ProGroup TM automated blood typing system also includes an LASO dot matrix printer 30 from Digi tal Equipment Corporation. The printer is connected to 4 would be reached after the user identified himself in the logon step 102 and was accepted as a permissible user. Processing would then vector to the main menu screen 104 wherein all the options shown in FIG. 3 65 would be presented to the user. If he selected option 100, processing would be vectored to the liquid han dling menu 106 after going through a logon step (not shown). The liquid handling menu selection allows the 4,873,633 10 liquid-handler. A set of six 96-well microplates is loaded into the plate stacker on the other side of the liquid-ham dler. Each plate can hold sample from eight donors. user to specify the process parameters of the liquid handling assay for blood typing, NTD determination and repeat liquid handling with or without bromelin pre-treatment. Since these process parameters are criti cal to accuracy of results, only supervisors or higher are Reagents for the test, including properly diluted cell suspension and antisera are placed in a row of wells in allowed to log into this function of the system. The user the reagent MicroTrof 62, along with a set of 12 tips. speci?es the following aspects of liquid-handling for the This MicroTrof also contains a row of reagents and a ABO/Rh test: whether or not a Bromelin pre-treatment is being row of tips for the antibody screening test to be de scribed later herein. The reagent MicroTrof is placed at 10 the back of the liquid-handler bed 74. used, choice 108 or 110, The predilution block 60 is placed just in front of the which blood typing test should be executed (the op reagent Microtrof. The block contains a trough of dilu tions are A, B, (A,B), Rh, Cell Suspension, A1, A2, B, ent, and several wells where the donor cells are mixed and Serm Blank), step 112 and, what test format is being with diluent. used (i.e., which tests are being carried out in which columns of the microplate to de?ne the format and ABO/Rh Testing Procedure arrangement of the template, i.e., the pattern of positive and negative reactions which de?ne the blood type), When the liquid-handler is set up and ready for oper ation the ?rst step is to carry out the automatic homing step 112. and priming procedure. Then the user pushes the The liquid handling may be done on the plates in any START key on the hand-held controller 94. The plate order and the plates may be read in any order. The stacker places a 96-well plate on the liquid-handler liquid handlers such as device 26 read the bar codes of table, and the bar code 59 on that plate is read. The bar all the tubes containing donor samples that are pipetted codes 21 on the ?rst group of eight tubes are also read. into a particular plate. These bar codes are sent to the Then the liquid-handler picks up the ?rst ProGroup host computer 20 along with the bar code of the plate into which samples from the identified tubes were 25 TM automated blood typing system tip from the carou~ sel and withdraws the appropriate amount of donor placed. The host computer knows which rows of each plasma from the top of the tube. Aliquots of plasma are plate contain aliquots of blood from each donor because placed, one by one into the wells of the plate 56 that are the control software of the liquid handler always places designated for reverse typing tests. Next the liquid-han samples in the plates in the same order relative to the dler withdraws the required quantity of donor red order of the tubes from which the samples were re blood cells for the forward typing tests. Aliquots of moved. The identi?cation data of the plates and the cells are placed into the wells of the predilution block WBNs that are in each plate are stored by the host 60, which already contain the required amount of dilu computer in a lookup table along with any tag data that ent. This diluent had been placed in the wells by the the liquid handler appends to the identi?cation data for the plate bar code. Such tag data can include data that 35 microprocessor of the liquid handler previously in ac the plate is a steps off center plate (this will be explained cordance with data received from the host regarding more fully below) or is a donor cell suspension or rea how much dilution the supervisor wants in the test procedure for red blood cells. The diluent and cells are mixed according to data regarding how many mixes that are desired which was speci?ed by the user and sent from the host to the liquid handler, and then the gent cell suspension plate etc. Because this data is per manently stored in a look up table, the order in which the plates are read by the plate reader is unimportant. The plates can be loaded in the plate reader in any order. The plate reader then reads the bar code for each plate and sends it to the host computer 20. The host computer then looks up the bar code in the look up table, and determines if any special instructions need to 45 tainer (not shown). When the microplate 56 is ?lled with donor samples, be sent back to the plate reader, e.g. as to wavelength to be used for the light or whether or not to do a steps off the l2-channel head picks up reagents from the reagent Micro-trof 62 and ?lls the row of the plate 56 containing the ?rst donor’s samples with the pattern of reagents center series of light absorbance readings. Parameters mixture is pipetted into the forward testing wells of the microplate. The used ProGroup TM automated blood typing system tip is then ejected into a used tip con 50 speci?ed by the user in the liquid handling procedure set-up step previously described (the user must ?ll the The user de?nes the test protocol by specifying in reagent trof 62 wells with the proper reagents in accor steps 114 and 116 for no bromelin and bromelin pre dance with the desired pattern). The row of Pro/Pette treatment respectively the: tips used for this operation is then ejected back into the volume of donor plasma to be pipetted into the wells 55 Microtrof. of the microplate, At this point, the liquid-handling on the plate is ?n volume of donor cells, the volume of diluent to be ished. The plate is loaded back into the plate stacker to mixed with the cells (thus the concentration of the await completion of the remaining ?ve plates in that donor cell suspension) and the number of mixes, group. concentration of the reagent cell suspension, and When all six are done, they are moved to the orbital titer of the reagent antisera. shaker for resuspension. Next they are centrifuged at Liquid Handler Set-Up Bar coded tubes containing properly prepared donor low speed to cause the cells to settle. Then, they are resuspended. This sequence of steps disperses the cells when the test results are negative, but causes the cells in mated blood typing system carousel. The carousel holds 65 a positive test to clump together in a tight button at the bottom of the well. 48 tubes and 48 ProGroup TM automated blood typing blood samples are loaded into the ProGroup TM auto system disposable tips. The carousel mounts at the side of the ProGroup TM automated blood typing system The ?nal step is to load each plate into the automatic plate reader. Plate reading can occur no less than three