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Optimization and Implementation of the Sysmex SP-1000i in a Multispecies Laboratory

Samantha Wildeboer, MT(ASCP), MSPfizer Global Research and Development

Groton, CT

Sysmex User GroupOctober 16, 2012

Outline

• Pfizer Groton Clinical Pathology Laboratory

• Former Process

• Evaluation

• Optimization– Initial Set-up

– Troubleshooting

• Implementation

• Project Outcomes

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Pfizer Drug Safety Research and DevelopmentGroton Clinical Pathology Laboratory

• Multispecies Clinical Pathology Laboratory Supporting:– Multispieces preclinical GLP and non-GLP safety studies

– Multi-site colony health monitoring

– Research and Business Unit exploratory studies

– Phase I/II Clinical Support

– Veterinary Medicine Research and Development

– Average two species, 50-100 samples/day

– Species: Monkey, Dog, Rat, Mouse, Rabbit, Cat, Pig, Horse, Cow, and Goat

• Platforms include:– Siemens:120/2120, 1800/2400, Centaur, Atlas, Immulite 2000,

CentraLink Middleware

– Stago Compact

– Sysmex SP-1000i, XT-2000iV

– Cerner Millennium LIS

Former Process –Hematology Sample Processing

• Pre-label slides with required information

• After Samples received, slides would be made manually– Large animal – average 30.36 seconds/slide

– Rodent – average 33.76 seconds/slide

– Average of 20 min/day (based on average sample number)

– 83.3 hours/year

• Samples sent to instrumentation for analysis

• Slides left to dry

• Load slide carousels and stain using a Wescor 7120 Aerospray

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Former Process – WescorEosin/Thiazin/Azure B/Methylene Blue Stain

Canine

Non-Human Primate• Advantages– Staining time

< 5 minutes

– Prepared reagents

• Disadvantages– 12 slide carousel

– Very little customization (Lighter/Darker)

– No staining of rabbit basophils(basophils in general)

– Pre-fix used for bone marrow staining

RabbitBasophils

Inspiration!

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Evaluation of Available Technology

• Gathered our requirements:– Stand alone– Read barcode– Interface to LIS (Cerner)– Print study important information on slide– Able to support our tube size(s)– Different settings for variable hematocrit– Several stain protocols available– Make a smear and stain, or just stain

• Contacted Vendors

Sysmex SP-1000i Highlights

• Stand alone

• Can interface with LIS

• Prints up to three lines (3D barcode available)

• No premixing of stain

• Makes smears, stains, or stains premade slides

• Supports 13 x 75 (2 mL) and Microtainer (500 uL)

• Variable angle settings (hematocrit)

• Multiple stain protocols available

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Optimization – Initial Settings

• On-site technical specialist provided:– Training– Evaluation of dog, non-human primate, and rat

samples– Smear angles– Initial optimization of available stain (Wright-

Giemsa)– pH testing?– NERL Water

Optimization – Initial Settings (cont.)

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Optimization – Initial Settings (cont.)

Optimization – The Next Steps

Stain Optimization

Date: __________________

Species/Slide #: _______________

Settings

Stain 1 prefix: ___________

Stain 1 Dry Time: ___________

Stain Cycle: ___________

Stain 1 Dilution: ___________

Stain 1 Dilution Time: ___________

Stain 2 Dilution Time: ___________

Rinse: ___________

Heater: ___________

Dry Time: ___________

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Optimization – Artifact Crenation

Low grade, appearing on 85% of slides

Optimization – Troubleshooting

• Probably not the stain

• Slide preparation or drying

• Slides (try a new lot)

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Optimization – May Grüenwald/Giemsa Stain

• Feedback to Sysmex– Contacted SP-1000i users in Germany

– Provided May Grüenwald/Giemsa staining protocol

• SP-1000i was converted to the two stain system– We loved the stain, but the crenation was still

present

Optimization - Troubleshooting

• Sysmex provided two SP-1000i Experts to help troubleshoot the issue– Back to a single stain system (PROTOCOL)– Determined to be a combination of things

o Different slide typeo Adjustment to the smear settings

Wright-Giemsa - Rodent

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The final battle: Wrights-Giemsa vs. May Grüenwald-Giemsa

Unanimous vote for May-Grüenwald!

May-Grüenwald-Giemsa - Rodent

May Grüenwald-Giemsa

Human

Non-Human Primate

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May Grüenwald-Giemsa

Canine

Rodent

Wescor Stain

Rabbit

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May Grüenwald-Giemsa

Rabbit

Basophils!

Implementation - Validation

• Acceptable Stain Quality

– 5 samples from four selected representative species (Human, Rat, Dog, and Monkey) will be stained and evaluated for acceptable stain quality.

– Cells should be easily identified, including Red Blood Cell and platelet morphology. Manual differentials performed will match automated differentials from the Siemens Advia 120 Hematology analyzer.

• Sample Identification

– At least 10 samples will be placed on the SP-1000iand slides prepared by the unit will be visually checked to ensure correct sample identification.

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Implementation - Metrics

• < 1 min to load the SP-1000i with 40 specimens

– Average of 0.825 min/day (based on average sample number)

– 3.4 hours/year

• Pre-labeling/post-labeling

• Stain Time Loss 5 min vs. 40 min (hands off)

• Shorter turn around to get the specimens to the hematology analyzer

• Approximately 80 Technologist hours saved/year!

Additional Utilization

• Bone marrow staining– Cytospin preps

– Histology brush preps

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Rat Bone Marrow Cells

SP-1000i Project Outcomes

• Improvement in stain quality

• Standardization of peripheral blood smears

• Efficiency gains

• Allows for utilization of cell imaging and classification technology (CellaVision)

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Questions?

Back-Up Slides

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May-Grüenwald-Giemsa Stain Protocol

Stain 1 prefix Use

Stain 1 Fix time 00’03

Stain 1 Dry time 00’05

Stain Cycle 5

Stain 1 Staining 04’30

Stain 1 dilution USE

Stain 1 dilution time 05’00

Stain 2 dilution time 20’00

Rinse Use

Heater Not use

Dry time 03’00

All pictures included in this presentation were taken using

CellaVision DM96

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The final battle: Wrights-Giemsa vs. May Grüenwald -Feline

The final battle: Wrights-Giemsa vs. May Grüenwald -Mouse

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The final battle: Wrights-Giemsa vs. May Grüenwald –Non-human Primate

The final battle: Wrights-Giemsa vs. May Grüenwald -Canine

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The final battle: Wrights-Giemsa vs. May Grüenwald -Rat