An Analysis of Parvo B 19 In Source Plasma Andrew Conrad Ph'D'

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An Analysis of Parvo B 19In Source PlasmaAndrew
Conrad Ph.D.
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INTRODUCTION

• We are currently testing for Parvo B 19 in an
  effort to remove the high titer samples from
  pools for manufacture.
• 40 of individual samples tested were IgG
  positive 75 of those had low level viremia.
• Removal of all Parvo B 19 is not feasible and
  would cause dramatic shortages of plasma.

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OBJECTIVE

• We developed a mechanism to eliminate the high
  titer, antibody negative, (Infectious?) Parvo B19
  samples from plasma pools for manufacture.

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Robotic Pooling Device
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Steps to Build Pool a (8x8x8)512 Member Pool
The 8 Tecan pipettes draw from all (colored) ROWS
in COLUMN Y1 in LAYER X1 and makes 3 deposits
into secondary pool vials A) 8 individual
aliquots into ROW (Z1-Z8colors) pool vials B)
all 8 pipettes aliquots into the single COLUMN
Y1 pool vial C) all 8 pipettes aliquot into the
single LAYER X1 pool vial
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Three Dimensional Plasma Pool Sample Matrix
8X8X8 (512)
X1
Z3
Y3
Primary Pool X1 Y3 Z3 Found Positive
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Three Dimensional Plasma Pool Matrix

• Master Pools are diluted 11000 (512,000 fold
  dilution) Sensitivity 20 copies per/Ml 10 6
• Result Negative
• conclude that all component samples are below
  the cutoff i.e. Not Implicated and will not
  cause high level contamination of the pool.
• Master Pool Result Positive
• resolve through testing of diluted primary
  pools and the implicated sample

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Resolution of the Individual Sample

• Primary pools must be diluted or quantitative
  analyzed or low titer samples will begin to
  confuse the resolution.
• Up to 30 of the samples could have low level
  viremia.
• Recommended cutoffs.

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Parvo Investigation Algorithm

• Retrieve samples of prior and subsequent bleeds
  for individual Parvo PCR analysis
• Determine Parvo antibody status
• Determine Parvo DNA levels over time
• Determine prevalence of the high titer samples.

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Composite Results
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Ideal Subject
determine viremic period and shape of curve for
each donor and examine antibody status (IgG vs.
IgM) throughout viremic window
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Summary of 20 donors
First appearance of IgM
First appearance of IgG
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Summary Points

• IgM Appears within 14 days of Viremia
• persists for several months
• IgG Appears after about 3 weeks
• Titer IgG remains elevated for years
• Viremia There is an immediate burst in viral
  load . The Average peak viral load is gt 1011
• Virus persists for years
• Viral Titers peak at about two weeks and
  immediately prior to IgM. Viral load then drops
  rapidly and remains at low levels for years.
  Highest titers can be above 10 10

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Incidence and Prevalence of Parvo B19

• Incidence of high titer infections 2002 was
  1/3281 donations. We feel this truly represents
  new infections because of the short duration of
  high viremia. (gt10 6 copies /ML)
• Prevalence of donors with some viremia is 1/3
• Breakdown according to season.

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2002 Incidence of Parvo B19 High Titer Samples