Leica Biosystems Bond Oracle HER2 IHC System - Manual (Page 19)

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Leica Biosystems Bond Oracle HER2 IHC System Instructions for Use TA9145 EN-CE-Rev_E 18/06/2013
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masked and assessed in a randomized fashion by a single trained observer to determine Lot-
to-Lot reproducibility.
An evaluation of the slides (tests and controls) from the lot-to-lot investigation indicated that
36/36 data points could be interpreted. No variation in staining occurred in the 36 data points
between the three different manufacturing lots of the Bond Oracle HER2 IHC System. Staining
with the Bond Oracle HER2 IHC System is consistent across manufacturing batches.
D. Between Laboratory Reproducibility
Between laboratory reproducibility testing of the Bond Oracle HER2 IHC System was evaluated
at 3 sites, Leica Biosystems Newcastle Ltd (Site A), and two independent laboratories
(Sites B and C) on a total of 192 sections from a TMA comprising of 20 invasive breast
tumors and 24 HER2 Control Slides. Of the 192 TMA sections stained, 96 were stained with
the HER2 Primary Antibody and 96 with the HER2 Negative Control reagent. All slides were
stained with the Bond Oracle HER2 IHC System on the BOND fully automated advanced
staining system. The slides were evaluated in 8 independent runs performed within each of
the 3 different investigational sites using a Bond Oracle HER2 IHC System from the same
manufacturing batch. Stained slides were blinded and assessed in a randomized fashion by
a single experienced observer at Leica Biosystems, Newcastle Ltd to determine between
laboratory reproducibility.
An evaluation of the slides from the between laboratory reproducibility investigation indicated
that 1477/1920 (76.93%) test data points could be interpreted. 443 test data points could not
be interpreted due to:
a) Inadequate performance of the HER2 Control slide on 2/24 occasions resulting in 2 runs/160
test data points being removed. This event occurred once at Site A and once at Site B (80 data
test points per investigational site removed).
b) Deviation from the test plan at Site C, in which 24 slides in total were manually
counterstained with hematoxylin following Bond Oracle HER2 IHC System staining. This
resulted in excessive counterstaining of both HER2 control slides and TMA test data points
resulting in 240 data points being removed.
c) Loss of invasive tumor resulting in 23 test data points being removed. This event occurred
on 23 occasions at Site A and was a direct result of loss of tissue in the TMA block on production
of the 192 consecutive TMA sections required to complete this investigation.
d) Uninterpretable staining due to inadequate washing by the BOND fully automated advanced
staining system resulting in 20 data points being removed.
An evaluation of the interpretable slides in the between laboratory precision investigation
indicated that variation in staining occurred 79 (5.28%) out of a possible 1477 staining events.
Of these, 14/1477 (0.95%) occasions represented variations from 0 to 1+ or 2+ to 3+ and as
such did not represent a change from clinically positive to clinically negative or vice versa in a
2x2 data assessment. Pass value = 99.05% (95% CI = 98.42% to 99.46%). Of the 14 staining
events, 5/1477 (0.34%) staining events occurred at Leica Biosystems, Newcastle, Ltd (Site A),
8/1477 (0.54%) occurred at Site B and 1/1477 (0.07%) occured at Site C.
The remaining 65/1477 (4.40%) staining events showed variation from 2+ to 1+ or 2+ to 0
and therefore would represent a change from clinically positive to clinically negative or vice
versa in a 2x2 data assessment. Pass value = 95.6% (95% CI = 94.42% to 96.54%). Of the
65 clinically significant changes, 11/65 (16.9%) occurred at Leica Biosystems, Newcastle,
Ltd (Site A), 24/65 (36.9%) occurred at Site B and 30/65 (46.1%) occured at Site C. Of the
clinically significant changes on no occasions did a 3+ change to a negative (0 or 1+) result
or vice versa.
E. Inter-Observer Reproducibility
40 randomly selected invasive breast cancer cases, providing an equal distribution of each of the
HER2 IHC grades (resection specimens) were consecutively sectioned and provided to Leica
Biosystems, Newcastle Ltd (Site A), Site B and Site C for staining and interpretation. The sections