# Presentation: 657815

## BD Solution for CD4 Testing

## CD4 Testing:  Safe-Simple-Reliable

## Absolute CD4 Counts

- _Accurate_ and _reliable_ counts of CD4+ T lymphocytes is essential in assessing immune status and managing healthcare of HIV infected patients
- _Accurate_ and _reliable_ results are dependent on
- Blood sample integrity
- Qualified reagents and sample processing protocol
- Validated instrument platform

## BD Technology Solutions

- Blood Specimen Collection
- BD Vacutainer® Brand offers a complete system of tubes, needles and needle holders, sharps collectors, and safety devices that are optimally designed and tested for controlling preanalytical variability throughout the specimen collection and handling process

- CD4 Test Platform
- BD FACSCount System is a complete solution: an automated instrument with dedicated reagent kit designed specifically for enumerating the absolute counts of CD4 and CD8 T lymphocytes

## Reliable results are dependent on a quality sample

## Laboratory Guidelines

**Universal Laboratory Precautions**
- Wear lab coats and gloves
- Never pipette by mouth – use safety pipetting devices
- Never re-cap needles – follow ‘needlestick’ prevention practices and use appropriate sharps collection devices
**BD Vacutainer Brand Blood Collection System**
- Recommend K3EDTA  anticoagulant evacuated tubes
- Spray-dried anticoagulent ensures no sample dilution for accurate absolute counts
- Guaranteed quality, training and technical support

## Sample Collection

## Sample Integrity

- Critical Guidelines
- Correct anticoagulent – K3EDTA – spray dried – no dilution for accurate absolute count
- Correct collection volume for proper anticoagulation
- Mix tube well after collection to prevent clotting
- Label appropriately as defined in Laboratory procedures
- Store and transport specimens at room temperature (64-72oF or 18-22oC)
- Transport speciment in appropriate leakproof container as per local regulations
- Perform test within 48hr (preferred) but no later than 72hrs after drawing blood specimen

## BD FACSCount SystemIntroduction

## Platform Simplicity

*Features*
- Easy-to-use
- Reliable
- Complete, stand-alone benchtop platform
- Consistent, accurate counts on samples with low CD4 values
- Automated QC, error conditions detected & flagged
- Absolute enumeration of CD4, CD8 and CD3 cells

*Benefits*
- Minimal operator training
- Low maintenance costs
- No special hook-up or computers required
- Facilitates effective management of HIV therapy
- Confidence in results
- Comprehensive test menu

## Instrument Design

## Instrument Specification

## Instrument Specification

## Operational Simplicity

- Complete system – easy to install and use
- Requires no external computer
- Minimal training for set-up and run samples
- Unique software algorithm identifies lymphocytes of interest
- Instrument QC verified using Control reagent kit
- Operates on standard electrical power

## System Software and Analysis

- FACSCount automatically acquires a fixed number of events
- Software evaluates the cluster of cells and beads using an eliptical orbital region that localises on the centre of each population
- Sufficient numbers of cells (upto 30,000) are collected to ensure statistically accurate results
- Fully automatic – no intervention
- If data meets QC criteria, results are reported as absolute counts

## Analysis and Reports

- Software provides automated report with no operator intervention
- Printed report quantifies CD4+, CD8+ and CD3+ T lymphocytes as absolute numbers per uL (mm3) of blood.  As well the CD4+/CD8+ cell ratio, and the average CD3+ cell count based on the average of CD3+ values from the same blood stained in the 2 sample tubes
- The reportable ranges for FACSCount are:
- CD4	1-2000 cells / L
- CD8	1-2000 cells / L
- CD3	1-3500 cells / L

## System Quality Control

- Controls should be run at beginning of each day by adding 4 Control bead populations to a control blood sample
- Control run verifies linearity of the system, and reports the correlation, slope and intercept of observed counts versus expected counts
- FACSCount QC is built-in to the software so erroneous results are not reported
- Results (Pass or Fail) are reported on each subsequent sample

## QC Analysis and Report

## FACSCount Accuracy

## FACSCount Precision

## Normal Reference Range

## Simple Sample Preparation Protocol

## Laboratory Guidelines

- Perform test within 48hrs (preferred), but no later than 72hr post collection
- Mix blood specimens gently before test
- Accurrate pipetting for reproducible results
- Vortex sample preps gently for adequate mixing
- Incubate samples in the dark
- Follow maunfacturers instructions
- Preferably analyse samples immediately after staining

## BD FACSCount Reagent Kit

- Reagents for 50 tests
- CD4/CD8/CD3 reagents in convenient pair sets with Tab for sample identification
- CD4/CD3 tube identified by green cap
- CD8/CD3 tube identified by clear cap
- Fixative solution – 2 x 5ml vials of 5% formaldehyde in PBS – for sample stability and biohazard protection
- Shelf-life – stable until date included on package
- Storage at 2-8oC (36-46oF)

## BD FACSCount Control Kit

- Provide 4 bead concentrations (zero, low, med, hi) to setup the instrument and check linearity
- 20 test runs per Control Kit in reagent tube pairs, with color coded tops
- Pair 1
- Zero –yellow top ( 0 beads / 50 L )
- Low – red top ( ~50 beads / 50 L )
- Pair 2
- Medium – blue top ( ~250 beads / 50 L )
- High – purple top (~1000 beads / 50 L )
- Shelf-life – stable until date included on package
- Storage at 2-8oC (36-46oF)

## Simple Whole Blood Sample Processing

## Sample Processing – Step by Step

## Sample Processing – Step by Step

## BD FACSCount Solution

- Simple

- Qualified Reagents

- Automated and
- Accurate Results