Manuscript received 09/04/96; accepted 10/15/96Laboratory Robotics and Automationq 1997 John Wiley & Sons, Inc. 0895-7533/97/010021-09 21

LRA, Vol. 9, pp. 21–29

Laboratory Robotics: AnAutomated Tool forPreparing IonChromatography CalibrationStandards

James L. Chadwick

This article describes the use of a laboratory robotas a tool to prepare multilevel calibration standardsfor several on-line ion chromatograph (IC) systems.The multichannel on-line IC systems were configuredto automate the sampling and analysis of the chem-istry in the tests. Analytical channels in the ICs wereset up for traditional anions and cations; therefore,calibration standards for anion and cation specieswould be required.

The use of the robot as a means to prepare cali-bration standards has provided significant advan-tages and benefits to the testing environment. Theseadvantages and benefits include the following:

1. accurate and precisely prepared calibrationstandards in individually capped containersalong with calibration level identification,

2. automated and unattended multispeciespreparation for both anion and cation analyticalchannels,

3. the ability to free up a test operator from arepetitive routine and reapply those efforts to testoperations, and

4. enables tracking and verification of each cal-ibration standard prepared to support laboratoryQA/QC plans.

System requirements and configuration, roboticoperations, operational checks, work force require-ments, analytical verification, accuracy and preci-

James L. Chadwick, Senior Scientist, Westinghouse Bettis, P.O.Box 79, West Mifflin, PA 15122.

sion of prepared solutions, and robotic downtime arediscussed in detail. q 1997 John Wiley & Sons, Inc.

BACKGROUND

Ion chromatography (IC) was introduced in 1975 bySmall, Stevens, and Baumann [1]. The technique de-veloped in a short time, from a new way of detectinga few inorganic anions to a versatile analytical tech-nique for ionic species of all kinds. Modern IC nowinvolves unique combinations of a number of sepa-ration systems with appropriate detectors. In 1983,Westinghouse Bettis [2] established a contract withDionex Corporation to develop a two-channel on-lineIC system. The base system was a 2120i IC controlledwith a Hewlett-Packard 9816 microcomputer. Thisautomation breakthrough led to additional on-line ICsystems over the next 3 years, bringing the total to 49on-line channels monitoring 37 test systems, manywith multiple sample locations. A variety of chem-istry matrices were being tested, necessitating theneed to formulate a single multilevel calibration stan-dard containing most of the same ionic species asused in each of the test facilities. High quantities offresh calibration standards were required to supportthe large number of separate on-line IC analyzers inoperation. The work force effort to support the IC cal-ibration requirements became extensive, leading toreduced test follow. To resolve this dilemma, a pro-posal to automate the IC standard preparation pro-cess by utilizing a laboratory robot was formulated.The proposal described how a robot would perform

22 Chadwick

the task of preparing accurate and precise multilevelIC calibration standards, yet reflect a work force sav-ings. In September 1989, Zymark installed at Bettis alaboratory robot to prepare mixed anionic and cation-ic species calibration standards to support continu-ous on-line IC operations.

JUSTIFICATION AND BENEFITS OF AROBOTICS SYSTEMTechnical justification to procure a laboratory robot-ics system was very straightforward. Hazardous ma-terials concerns or environmental effects were not anissue with this application. The primary advantagesderived as a result of installing a robotics system forstandards preparation are listed in the following:

● The robot has the ability to operate unattendedor on back shifts. In doing so, a skilled technicianhas been relieved of a boring repetitive job tomake more efficient use of time for test prepara-tion and test follow.

● The problem of technicians’ busy schedules in-terfering with preparation of standard solutionshas been eliminated. QA/QC requirements werejeopardized because calibration of equipmentcould not be maintained without fresh calibra-tion solutions.

● The quality of calibration standard has im-proved by eliminating gross error, occasionallyencountered in human prepared standards (i.e.,missing ionic species, several-fold error in con-centration).

● With each standard solution prepared, a recordof its contents is printed for QA/QC traceability.The robot also maintains record of analytical ver-ification for each standard.

DEFINING THE ROBOT OPERATINGPROCEDUREOnce justification for need was established, the ques-tion of robot capability required investigation. Thesystem would be required to prepare standard solu-tions with precision and accuracy equivalent to thatof a technician or scientist.

The ideal role of the robot would be to preparemultilevel mixed anion/cation calibration standardsin volumes that are easy to handle by human androbot. The system must be capable of maintaining re-cords of the contents, final dilution, and analyticalresults of each prepared standard. In addition, thesystem must be capable of either flagging the operator

of an improperly prepared solution or discarding orisolating it for proper disposal.

To accomplish this, a step-by-step list of actionsor laboratory unit operations (LUOs) [3] had to becreated to determine exactly what the robot could orcould not do based on the ideal role described above.At this time, operational checks and calculationswere not considered because they would be ad-dressed during debugging. To determine a list ofLUOs required for this operation, a flowchart wasprepared to show exactly how a standard solutionwould be prepared as if it were being prepared by aperson. This also helped in determining if precisionand accuracy requirements could be met. The labo-ratory procedure of preparing IC calibration stan-dards would then be systematically translated [4]into terms of

● the required LUO,

● their sequence in the procedure, and

● what happens in each LUO.

Listed in the following are the LUOs based onman-made standard preparation.

● Manipulation (fleakers)

● Liquid Handling (add water and preservativeto fleaker)

● Conditioning (stirring)

● Manipulation (weigh dish on balance pan)

● Weighing (weigh powder)

● Manipulation (powder dumped into fleaker)

● Conditioning (stir fleaker to mix)

● Liquid Handling (add reagents and final dilu-tion)

● Conditioning (stir to mix)

● Manipulation (move fleaker to IC sampler)

● Measurement (IC analysis)

● Manipulation (return fleaker to storage)

● Documentation (standard identification andanalysis results stored)

This LUO list along with detailed documentation onadditional required robotics operations was sent toZymark Corporation for logic review and parts avail-ability. Zymark replied favorably with respect tooff-the-shelf hardware, pysections, and equipment. Apysection is robotic hardware integrally designed toperform some LUO and is attached to key referencepoints on a circular mounting plate. All standard op-erating commands for each pysection are prepro-grammed on an accompanying disc using Zymark’sEASYLAB software.

Laboratory Robotics: An Automated Tool for Preparing Ion Chromatography Calibration Standards 23

There were two sections that required custommanufacture and fit. This meant the final systemwould have to be set up and tested at the vendor’sassembly facility prior to shipment. Due to Westing-house Bettis policies, only the custom pysectionscould be tested and debugged at Zymark. The re-maining setup and testing was completed on site byZymark without significant problem.

Special Designed Pysections

Two robot pysections were designed, assembled, andtop-level programmed in EASYLAB pytechnologycontrol language specifically for this robotic appli-cation. First, a bar-code reading station was designedto read a bar-code label placed on the side of a fleaker.The bar-code label is a single six-digit number placedon the side of a fleaker that the scanner and robotcontroller interprets as (1) a standard identificationused for traceability and (2) an identifier that directsthe robot to what calibration level to prepare. Thenumbers range between 100000 and 699999, incre-mented singly. Each incremental range of 100000 re-lates to a calibration level (i.e., 100000 through199999 for level one, continuing respectively up to699999). The section was designed to contain a ro-tating platform and a 1.0 mw class IV laser scanner.The platform rotates a fleaker (placed there by therobot), enabling the bar-code label to be scanned bythe laser reader. Once the label is read and informa-tion is stored in the controller, the fleaker is removedfrom the platform.

The second is a variable speed magnetic stirringsection. This section was designed to mix and dis-solve all additions to the 300 mL fleaker. Both pysec-tions would be controlled through relays and powerswitches from a Power and Event Controller (PEC).

Original Robotics System

The original robotics system required several piecesof additional equipment not available from Zymark.The following is a list of that additional equipment:

Balances—Mettler AE200 with communica-tions interface

Mettler PM400 with communications inter-face

Bar-code scanner/reader—Symbols Technol-ogies

Fleakers and caps—Caps from Zymark

Test tubes—30 2 150 mm

Vials with caps—30 2 150 mm

Reservoirs—1 and 2 L plastic

The pysections originally procured have remainedunchanged. Upgrades to the robot arm, controller,and computer system have been installed severaltimes since 1989. The following is a list of pysectionsfrom the original system that are still in use today:

Fleaker rack (2), 300 mL size

Bar-code reader

Balance section (PM400)*

Magnetic stirrer

Test tube rack (50 2 150 mm size tubes)

Cap/uncap station (scheduled for upgrade)

Balance section (AE200)

Waste chute

Hand C

Hand G

Hand E

Master lab station (2)

Power and event controller (2)

Zymate II robot arm

Controller, disc drive, and monitor

This system operated until the System V controllerand PC computer interface was introduced about 2years later.

System Upgrades to Date

The program and dictionaries to operate this systemrequired over half the available memory of the Zy-mate II controller. The first major upgrade after about2 years of operation was to the System V controllerand PC interface. This improved process speed andmade available additional memory.

The second major upgrade, in December 1993,was the addition of a new PC interface system andhigh-speed XP Arm. This made a difference of 30%less in preparation time of a single solution. Arm-position calibrations are maintained longer, and fre-quency of routine adjustments have been reducedfrom monthly to quarterly. This upgrade also re-sulted in a reduction of preparation errors by at least50%.

The last upgrade included the addition of Con-current programming capabilities. This enabled non-robotic operations (i.e., liquid dispensing, weighing,

*One balance section was modified to mount an air operated armholding six liquid delivery lines that would swing over the fleaker(while on the balance) and deliver predetermined volumes of liq-uid. It also contains a small funnel attached to a drain that allowsflushing of the lines while the arm is in the rest or retracted po-sition.

24 Chadwick

Figure 1. Current robot config-uration. The PC system andprinter are located on an adja-cent bench top.

or any operation not using the robot arm) to be per-formed simultaneously with robotic functions. A Di-onex DX-100 single-channel (anionic) IC system wasadded to support the actual analysis of every stan-dard solution prepared by the robot. It should benoted that reagents were chosen to utilize both theanion and cation in the standard matrix. For this rea-son, it was determined that only anion checks of theprepared standards were needed. A Zymark LC/ICSipping Station was also added to support the IC sys-tem. This final and current robot configuration isshown in Figure 1 as an overhead outline layout.

DETAILED SYSTEM OPERATIONAfter determining the proper robotic LUOs, the sys-tem programming was assembled in modules. Mostpysection command modules (i.e., Get From Rack. 5)required some modification for smooth and efficientoperation. The operating procedure to prepare singleor multiple IC standard solutions was broken intotwo steps: Manual Operations and Robotic Opera-tions.

Manual OperationThe following must be performed prior to each robotoperating session—corrections to each operation areto be completed only if necessary:

● the level of solution containers must be .1/3full to insure adequate supply for the operatingsession;● the level of dry chemical in each vial must be1/4 to 1/2 in. deep to allow for disbursement fromthe test tube;● all fleakers that have been bar-code labeled arecapped;● check to ensure a magnetic stirrer is in eachfleaker so that mixing of all reagents is complete;● check that at least 75 mL of ,14-day-old 4%formaldehyde solution is available so that thecorrect amount of fresh preservative is added;● a full rack of 10 clean empty test tubes is avail-able;● sufficient quantity of pipette tips for at leasttwice as many standards being prepared is avail-able;● the printer is on-line and gas pressure is at25–30 psi to prevent malfunction of air-operatedcomponents; and● power on the robot computer controller, ICcomputer, and Dionex IC.

Robotic OperationsCommencement of operations was designed to use asingle command. By typing ROBERTA and answer-

Laboratory Robotics: An Automated Tool for Preparing Ion Chromatography Calibration Standards 25

ing all remaining questions, using the computer key-board, the robot would begin preparing calibrationsolutions. The following is the current step-by-steprobot procedure to prepare a single IC calibrationstandard:

● Uncap the first fleaker in line and store the capin the holder.

● Remove the fleaker from the rack, and place iton the turntable for the bar-code reader.

● The fleaker is rotated several times; the laserscans the bar code and reads the number.

● The fleaker is transferred to the PM400 balancefor the addition of 150 g of DI water. During theaddition of water, the robot’s hands are switched,and a 0.2 mL aliquot of 4% formaldehyde istransferred to the fleaker. Hands are changedback, and the robot arm waits until the fleaker hasreceived the 150 g of water before continuing.

● The fleaker is transferred to the magnetic stir-ring bar section to start mixing.

● The robot changes hands to pick up an un-capped glass vial from RACK.3 and places it inthe AE200 balance.

● The robot then picks up the capped vial con-taining the dry chemical species from the samerack.

● The vial is uncapped and the cap stored on oneof the storage pedestals on the pysection.

● The robot adjusts the tube from a picking upgrip to a pouring grip and moves over top of thetest tube in the AE200 balance.

● Based on the bar-code numbering sequence,the controller has stored the target value of pow-der to add to the tube. The robot now pours fromthe tube using the vibration option in the handto slowly pour powder into the test tube. The bal-ance continually takes weight readings until theactual weight is within `/111% of the targetweight.

● Once the target weight has been satisfied, thepowder is transferred into the stirring fleaker.

● The robot changes hands and transfers thefleaker to the PM400 for addition of remainingionic species and final dilution water.

● The ionic species are added by a Master LabStation using 5 and 10 mL syringes. (Note thatthe volumes added are dependent upon the `/1 difference of the actual powder weight in re-lation to the targeted weight. The difference isautomatically calculated and adjustments aremade in all remaining additions so that all stan-

dards of the same level are prepared to the sameconcentration.) Frequently, final volumes differas much as 75 mL.

● The fleaker is transferred back to the stirrer forfinal mix.

● The fleaker is transferred to the IC sipping sta-tion where a sample is drawn into the IC via amaster lab syringe. The IC then analyzes the pre-pared solution.

● The fleaker is returned to its original positionalong with its cap.

● The robot then moves onto the next fleaker,provided one exists. If not, the arm parks thehand, and the system rests in an idle position.

PRECISION OF PREPAREDCALIBRATION SOLUTIONS

Figures 2 thru 6 are graphs reflecting the results of agroup of standards analyzed by the DX-100 IC. Eachgraph reflects the precision of a species from one runto another. It is important to note that the precisionof the IC analysis has not been factored out of thesegraphs. The results shown give a more realistic num-ber to the precision of the overall analytical resultsof actual test samples. Calibration of the DX-100 ICwas performed each time the robot ran using dilutedNIST anion standards.

Built-in Operational Checksand Calculations

During the procedure, checks to insure proper sys-tematic operation and to prevent inadvertent mal-functions had to be programmed into its operation.Any error in the preparation of a standard will jeop-ardize the validity of the robot operation along withthe IC analytical method.

Bar-Code Reader CheckThe bar-code reader makes three attempts at readingthe applied bar code before ending the operationwith an error message and exiting from the program.Clear and legible bar codes are required to preventimproper identification of a standard level.

Water Addition TimerDue to an unidentifiable communications error be-tween the PM400 balance and the System V control-ler, water over additions to the extent of flooding thebalance have occurred, leaving the balance inopera-ble. A programming timer was built into the mainprogram to halt dispensing of water after a predeter-

26 Chadwick

Figure 2. Species “A” concentration.

Figure 3. Species “B” concentration.

Laboratory Robotics: An Automated Tool for Preparing Ion Chromatography Calibration Standards 27

Figure 4. Species “C” concentration.

Figure 5. Species “D” concentration.

28 Chadwick

Figure 6. Dry species concentration.

mined length of time. This enables the robot to con-tinue preparing the standard. The robot automati-cally takes into account the water over addition andadjusts all further additions accordingly. This alsoeliminates possible damage to equipment due toflooding.

Powder Weighing ChecksThe powder is weighed into test tubes to a targetvalue based on the calibration level (bar-code num-ber). If the powder weight is .11% of the targetvalue, the total adjusted volume in the fleaker wouldexceed the maximum volume of the fleaker as wellas the range of the PM400 balance. In such a case,the robot will discard the test tube and its contentsto waste. This check is in the event of over additionof powder to the test tubes during the weighing pro-cess.

A second weight check is made when the powderis actually poured into the fleaker. A weight readingis taken after the powder is transferred to the test tubeand is recorded as a tare weight. A second reading istaken on the test tube after the powder has been trans-ferred to the fleaker. The difference from the secondto the first is the actual weight of powder added. It is

this actual weight that is used as a basis in determin-ing volumes of other species added and the final di-lution volume of DI water. If this weight exceeds thetarget range by .11%, the fleaker is returned to thestorage rack, and no other ionic species are added.

Standard IC Analysis CheckA Dionex DX-100 single-channel IC system checkseach standard prepared for anionic contents. The re-sults of these analyses are stored in the Dionex AI-450 software for future reference, in addition to ahard-copy chromatogram that can be used to visuallyinspect the chromatography of each prepared stan-dard. The robot prints out a summary report showingthe exact volumes of stock solutions added to eachfleaker. By using both IC results and a robot summaryreport, verification of the prepared standards can beeasily made. These results are stored permanentlyper laboratory QA/QC requirements.

STANDARD TRACKING ANDRECORD KEEPINGAll prepared calibration standards are controlled bybar-code number and preparation date. Two separate

Laboratory Robotics: An Automated Tool for Preparing Ion Chromatography Calibration Standards 29

reports are generated during each run of standards.One report lists the mass contents of each standardalong with the operator’s name and date of prepara-tion. The second report is the IC results of each pre-pared standard solution. The two reports are coor-dinated by prewritten IC schedule files that matchthe order in which the numbered fleakers are placedin the storage rack.

The robot report displays the volume of each offour liquid ionic additions, weight of powder added,and total volume of water added during preparationfor each solution. This information combined withthe preparation date, operator’s name, and identifi-cation gives the operator all the information neededto correctly evaluate the standard preparation. TheIC report is a verification that the solutions were pre-pared properly and a means to monitor deviationsthat occur during solution preparation.

ROBOTIC, DOWNTIME, SERVICE, ANDMAINTENANCEThe robot is covered by a service contract with Zym-ark Corporation for quarterly preventative mainte-nance (PM) visits and repair or full replacement ofall malfunctioning components. Unexpected failuresrequiring a service call have occurred on the averageof twice per year. Minor repairs not affecting qualityof calibration solution preparation are delayed untila PM visit. The frequency of PM visits has ensured avery dependable system supporting continuous on-line IC operations.

The DX-100 IC system is serviced separately byDionex Corporation with a PM schedule of five visitsper year.

Balances are cleaned and calibrated quarterly bya qualified vendor.

Overall, daily workforce support is very mini-mal. The primary operator effort required to preparethe robot for operation is mostly used for cleaningfleakers and preparing them for use. This is a dailyrequirement and usually is completed in about 15minutes for a run of 12–14 standards. Every othermonth the dry powder is replaced with fresh reagentgrade material, and the solution reservoirs are re-filled. The pipet tip rack is refilled monthly.

SUMMARYThe robot has proven to be a vital asset to the testlaboratory. Its reliability of operation has never been

a problem. The quality of standards prepared is re-flected in the high quality of on-line IC data produceddaily. It has enabled an increase in test productivityover the 5 years of operation by saving and redirect-ing a total of about 1.2 years.

The robot was relocated in May 1994 to a per-manent private room so its environment can be bettercontrolled and monitored. The future of this systemis promising. Plans are to keep it operational daily,well into the twenty-first century.

NOTICE

This report was prepared as an account of work spon-sored by the U.S. Government. Neither the UnitedStates, nor the U.S. Department of Energy, nor theU.S. Navy, nor any of their employees, nor any oftheir contractors, subcontractors, or their employees,make any warranty, express or implied, or assumeany legal liability or responsibility for the accuracy,completeness or usefulness of any information, ap-paratus, product, or process disclosed, or representthat its use would not infringe on privately ownedrights.

ACKNOWLEDGMENTS

A special thanks and appreciation are extended toJack Carr, Westinghouse Bettis, Ron Briggs, ZymarkCorporation, and Dale McBride, Dionex Corporation.

REFERENCES

[1] Joachim Weiss, Germany, “Handbook of Ion Chro-matography,” Dionex Corporation, Sunnyvale, CA.

[2] Gary Lynch, “Practical Experience with On-LineChromatography,” Process Control and Quality, Vol.1, Elsevier Science Publishers B. V., Amsterdam,249–263 (1991).

[3] Zymark Corporation, “Laboratory Robotics Hand-book,” Zymark Corporation, Hopkinton, MA, pp. 18,20 (1988).

[4] Zymark Corporation, “Laboratory Robotics Hand-book,” Zymark Corporation, Hopkinton, MA, p. 20(1988).