Innovative, Multi-platform, Affordable CD4+

Enumeration Reagents that Need No Cold Chain:

Impacting HIV/AIDS Management in Resource Poor
Settings

Dr.Anand Sivaraman,
ReaMetrix India Pvt. Ltd.
Bangalore 560092

© 2008 Reametrix, Inc. All rights

 HIV in India – Current Status
HIV Prevalence rate in India 0.36% (source, NACO 2007 Bulletin)
2.5 Million HIV Infections

Management of HIV Infection

Providing Anti Retroviral Therapy (ART)
Immunophenotyping
CD4 T-Lymphocyte Count
Viral Load Assay
PCR – quantitate virus levels in blood

Role of CD4+ T-Lymphocyte Count in HIV pathogenesis

Assessing immune status of patients
Indicating initiation of prophylactic ART therapy
Assessing patient response to therapy

© 2008 Reametrix, Inc. All rights 2

Current Issues with CD4 testing in Resource Poor
Settings ReaMetrix APPROACH
ISSUES
High cost of testing Undertook a study of the anatomy of
cost of CD4 testing
Poorly developed cold chain in the Worked on a locally relevant
transportation, storage & use of innovative product delivery model –
reagents dried reagents

Aging of blood samples that are Re-designed reagents to enable

transported to centralized testing
facilities
Complicated assay workflow with
multiple critical pipetting steps
leading to errors in CD4 count
enumeration
staining in collection centres
Simplified assay workflow with a
single critical pipetting step – reduced
chance of human error

© 2008 Reametrix, Inc. All rights 3

Clinical Information Generation – Current Scenario
Sample journey and cost accumulation
Provide
Acquire Transport Process Run
Results

Collection Centralized
centre Path Lab
Path Lab
Lab Infrastructure
Doctor
Capital Overheads
Hospital Tech Labor + Equipment
Amortization Layers of
Reagent
Cold margin
Syringe/Vacutaine chain/Courier at every
r/Overheads
step

• CD4 test example:

• ~2 to 5 times cost addition from reagent manufacture to cost to patient
• Average cycle time for results ~ 48 to 72 hours

© 2008 Reametrix, Inc. All rights 4

Innovations that helped reduce the cost of CD4 testing
Optimized the manufacturing processes to
enable a two-fold increase in antibody-
conjugate yield

Validated and standardized the use of open

source bright green and red dyes
Atto 488 (FITC equivalent)
Phycoerythrin-Dyomics 649 (PE-Cy5
equivalent)
Development of Dried Reagents
Uses proprietary stabilizers in formulation
Removes costs associated with
maintaining cold chain in storage and
transport

© 2008 Reametrix, Inc. All rights 5

 Innovation in Product Delivery : Issue of Aging of Blood samples

 In India, the real challenge lies in transporting blood samples from remote areas to
centralized testing centers.
 Often times, this leads to “aged” blood samples (> 48 hrs), making it unusable
 Blood collection centres need access to cold-chain to enable storage of liquid
reagents, if they were to stain samples and then ship them

CD3 capping
© 2008 Reametrix, Inc. All rights 6
 Unitized (one test per tube) dried-down format :
Eliminating the Issue of Aged Samples

Dried reagents can be stored and transported at room temperature

Blood can be collected, stained and fixed at the point of collection before being
shipped to a central testing facility at room temperature
The stained samples can be stored for up to 7 days at room temperate without any
statistical difference in CD4/CD8 counts on 1st day vs. the 7th day.

© 2008 Reametrix, Inc. All rights 7

 Simplification of Assay Workflow

Methodology
Employ simple Lyse/no wash protocol - single platform
technology

Unitized Format
Pre-dispensed reagents in unitized form provided as
ready to use tubes
Convenience & Simplified Work Flow
Just 2 pipetting steps – no separate pipetting of beads,
reagents etc.

© 2008 Reametrix, Inc. All rights 8

ReaMetrix HIV Reagents Performance Assessment
*
* *

* ­ values in cells per uL © 2008 Reametrix, Inc. All rights 9

 Building in Robustness in Reagents: Shelf Life exceeds 1 year

 Dried Reagents are stable at room

temperature for more than 12 months

 Enables use of dried reagents in resource

poor settings without cold chain/
refrigeration – enables increased
penetration into remote areas.
Stability of ReaMetrix Dried
Reagents at Room Temperature : do
not require Cold storage

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ReaMetrix Reagents :
Making the difference : Changing the Paradigm

All reagents already formulated, unitized and

dried
Steps as simple as mix & read
No cold chain in transportation or storage
Long shelf life and reduced waste
Less opportunity for pipetting errors – with fixed
volume pipettes, sample processing is made very
easy
All this without compromising quality or
affordability.
© 2008 Reametrix, Inc. All rights 11
In the words of the National Microbiology Reference Lab, Zimbabwe

© 2008 Reametrix, Inc. All rights 12

Key Centres who have already Validated Reagents
and Published Data from Validation
National AIDS Research Institute (NARI), Pune
Public Health Agency of Canada, Canada
National Microbiology Reference Lab, Zimbabwe and CDC,
Zimbabwe

© 2008 Reametrix, Inc. All rights 13

Publications & Presentations
1. Manian, BS, et al, Affordable Diagnostics — Changing the Paradigm in India,
Cytometry Part B (Clinical Cytometry) 74B (Suppl. 1):S117–S122 (2008)

2. Bhavani Venkatkrishnan, Pratibha Malini, K.L.Ravikumar, Anand Sivaraman,

Comparative Evaluation of an Indigenous CD4 Reagent Kit in a Dry Format
Requiring no Cold Chain, Proceedings of the International Conference on
Fundamental and Translational Research on HIV/AIDS, Mumbai, October 2008.

3. Rajesh Anand, Jammi Narasimham, Bhavani Venkatakrishnan, Wilson D'Souza, Sowmya

Narayanan, Donny Thomas, Anand Sivaraman, Multi-platform, Affordable CD4
Enumeration Reagents that need no Cold Chain: Enabling evidence-based
Management of HIV/AIDS in Resource-poor Settings, Proceedings of the
Association of Pathologists and Microbiologists (APCON), Chennai, December 2008.

© 2008 Reametrix, Inc. All rights 14