David R. Rank et al.

Patent Trials and Appeals Board

Oct 17, 2019

14318324 - (D) (P.T.A.B. Oct. 17, 2019)

UNITED STATES PATENT AND TRADEMARK OFFICE
UNITED STATES DEPARTMENT OF COMMERCE
United States Patent and Trademark Office
Address: COMMISSIONER FOR PATENTS
P.O. Box 1450
Alexandria, Virginia 22313-1450
www.uspto.gov
APPLICATION NO. FILING DATE FIRST NAMED INVENTOR ATTORNEY DOCKET NO. CONFIRMATION NO.
14/318,324 06/27/2014 David R. Rank 105-001108US 5505
57770 7590 10/17/2019
Pacific Biosciences of California, Inc.
1305 O' Brien Drive
Menlo Park, CA 94025
EXAMINER
GROSS, CHRISTOPHER M
ART UNIT PAPER NUMBER
1639
NOTIFICATION DATE DELIVERY MODE
10/17/2019 ELECTRONIC
Please find below and/or attached an Office communication concerning this application or proceeding.
The time period for reply, if any, is set in the attached communication.
Notice of the Office communication was sent electronically on above-indicated "Notification Date" to the
following e-mail address(es):
PATENTS@PACB.COM
PTOL-90A (Rev. 04/07)

UNITED STATES PATENT AND TRADEMARK OFFICE
____________

BEFORE THE PATENT TRIAL AND APPEAL BOARD
____________

Ex parte DAVID R. RANK, JEFFERY WEGENER, JONAS KORLACH,
DANIEL ROITMAN, YUE XU, JOHN LYLE, STEPHEN TURNER,
PAUL PELUSO, GEOFF OTTO, and RONALD L. CICERO
____________

Appeal 2018-001129
Application 14/318,324
Technology Center 1600
____________

Before ULRIKE W. JENKS, RICHARD J. SMITH, and DAVID COTTA,
Administrative Patent Judges.

COTTA, Administrative Patent Judge.

DECISION ON APPEAL
Pursuant to 35 U.S.C. § 134(a) Appellant1 appeals from the
Examiner’s decision to reject claims 1–13. We have jurisdiction under 35
U.S.C. § 6(b).
We reverse.

1 We use the word “Appellant” to refer to “applicant” as defined in 37
C.F.R. § 1.42. Appellant identifies the real party in interest as Pacific
Biosciences of California, Inc. App. Br. 2.

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STATEMENT OF THE CASE
The Specification discloses that “[t]here are a wide range of analytical
operations that may benefit from the ability to analyze the reaction of
individual molecules, relatively small numbers of molecules, or molecules at
relatively low concentrations.” Spec. ¶ 4. The Specification further
discloses that a number of approaches have been described for analyzing
reactions involving small numbers of molecules including by using “optical
confinement techniques . . . to ascertain signal information only from a
relatively small number of reactions, e.g., a single molecule, within an
optically confined area.” Id. ¶ 5. One method of optical confinement is to
use a zero mode waveguide (“ZMW”). Id. ¶ 15.
ZMWs are “generally characterized by the existence of a core
surrounded by a cladding, where the core is dimensioned such that it
precludes a substantial amount of electromagnetic radiation that is above a
cut-off frequency from propagating through the core.” Id. ¶ 56. “As a
result, when illuminated with light of a frequency below the cutoff
frequency, the light will only penetrate a short distance into the core,
effectively illuminating only a small fraction of the core’s volume.” Id. The
Specification discloses that reactions outside the illuminated portion of a
ZMW may “impact the reaction of interest or the monitoring of that reaction,
by affecting reaction kinetics through depletion of reagents, increasing
concentration of products, [or] contributing to signal background noise
levels.” Id. ¶ 58. The Specification thus teaches that “it may be further
desirable that reactions of interest be reduced or even eliminated from other
regions outside of the observation volume, e.g., on the overall substrate
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housing ZMWs, the cladding layer, etc., both inside and outside of the
observation volume.” Id.
The Specification discloses that the present invention is directed to
“methods for localizing individual molecules within a particular space or
volume, such that the spatial individuality of the molecule may be exploited,
e.g., chemically, optically, electrically, or the like.” Id. ¶ 54. “[S]elective
and preferential deposition and/or immobilization of the reaction
components within the observation volume [of a ZMW] are particular
advantages of the invention.” Id. ¶ 58.
Claims 1–13 are on appeal. Claim 1 is illustrative and reads as
follows:
1. A system comprising:
an array of nanoscale wells disposed through an opaque
layer, each well having a bottom surface formed from a
transparent layer underlying the opaque layer; wherein a
plurality of the wells in the array comprise an enzyme bound to
the bottom surface and located in an observation region; and
wherein portions of the array other than the bottom surface of
the wells are substantially free of the enzyme;
an illumination source that provides illumination to the
observation regions through the transparent layer; and
an optical detector that receives electromagnetic radiation
emitted through the transparent layer from the observation
regions.

Claim App’x 1.
The claims stand rejected as follows:
Claims 1–4 and 6–9 were rejected under 35 U.S.C. § 102(b) as
anticipated by Levene.2

2 Levene et al., Zero-Mode Waveguides for Single-Molecule Analysis at
High Concentrations, 299 SCIENCE 682–86 (2003) (“Levene”).
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Claims 1–9 and 11 were rejected under 35 U.S.C. § 103(a) as
obvious over the combination of Levene and Hofer.3
Claims 1–13 were rejected under 35 U.S.C. § 103(a) as obvious
over the combination of Levene, Hofer, and Sluka.4
Claims 1, 6, and 7 were rejected on the ground of non-statutory
obviousness-type double patenting over US Patent No. 7,302,146
(“the ’146 patent”) in view of Levene.
ANTICIPATION
Levene discloses a “zero-mode waveguide [ZMW], for highly
efficient single-molecule analysis at high fluorophore concentrations and its
application to enzyme analysis.” Levene 683. This is depicted in Figure 1
(reproduced below).

Figure 1 of Levene depicts “[a]n apparatus for single-molecule enzymology
using zero-mode waveguides.” Id. Levene’s ZMWs “were manufactured as
small holes in an 89-nm thick film of aluminum on fused silica coverslips.”
Id. at 684. Levene discloses that enzymes, like polymerase, can be adsorbed

3 Hofer et al., US Patent Publication No. 2003/0186914 A1, published Oct.
2, 2003 (“Hofer”).
4 Sluka et al., US Patent No. 5,851,840, issued Dec. 22, 1998 (“Sluka”).
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onto the bottom of the waveguide to allow direct observation of single-
molecule enzymatic activity. Id. at 683, 685. The supporting materials for
Levene disclose that “the enzyme bound nonspecifically to . . . fused silica”
and that “[a]fter immobilization, unbound enzyme was washed away by
extensive flushing with polymerization buffer.” Levene, Supplementary
Materials, http://science.sciencemag.org/content/sci/suppl/2003/01/29/299.
5607.682. DC1/Levene.SOM.pdf, (2003).
The issue presented in this appeal is whether the preponderance of the
evidence supports the Examiner’s determination that the portions of
Levene’s waveguides, other than the bottom surface of the wells, are
“substantially free” of the enzyme.
In finding Levene anticipated claims 1–4 and 6–9, the Examiner
concluded that polymerase bound to the silica substrate at the bottom of the
Levene’s ZMWs but did not bind to (or was washed from) the aluminum
that surrounds and forms the walls of Levene’s ZMWs, thus meeting the
requirement of claim 1 that the “portions of the array other than the bottom
surface of the wells are substantially free of the enzyme.” Ans. 2, 10. As
support, the Examiner points to the disclosure in Levene’s supporting
materials section that “unbound polymerase [excess] was washed away with
extensive flushing.” Id. at 10 (alteration in original). The Examiner points
out that this interpretation is consistent with Figure 1 of Levene (reproduced
above), which “plainly depicts . . . one enzyme . . . molecule at the bottom of
the observation region with none elsewhere.” Id. at 8.
Appellant argues that polymerase was present not just on the silica
bottoms of Levene’s ZMWs, but also on the aluminum surfaces that
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surround the ZMWs. App. Br. 10–12. Appellant relies on Korlach5 as
support. According to Appellant, Korlach discloses that polymerase binds to
untreated aluminum, even after it is washed to remove unbound polymerase.
Id. at 10–11. As further support, Appellant relies on the testimony of Dr.
Popovich that “[t]he washing conditions [described in Levene] would . . . not
disrupt nonspecific adsorption of the Sequenase enzyme elsewhere on the
ZMW array.” Popovich Decl.6 3. Dr. Popovich explains that “the washing
conditions [in Levene] did not disrupt the desired nonspecific adsorption of
the Sequenase enzyme to the bottom of the ZMWs” and thus “would also
not disrupt nonspecific adsorption of the Sequenase enzyme elsewhere on
the ZMW array.” Id.
Appellant argues that binding of polymerase to the aluminum surfaces
surrounding Levene’s ZMW’s is consistent with Levene’s teaching that its
ZMWs can be used to detect the activity of single molecule kinetics because
Levene speaks only to the presence or absence of enzyme in the observation
volume of the ZMW. Appellant explains:
Because illumination is effectively limited to a tiny observation
volume at the bottom of the ZMW and fluorescence is
effectively observed only from this observation volume,
activity of a single polymerase molecule can be monitored as it
incorporates fluorescently labeled nucleotides one at a time,

5 Korlach et al., Selective Aluminum Passivation for Targeted
Immobilization of Single DNA Polymerase Molecules in Zero-Mode
Waveguide Nanostructures, 105 PROC. NATL. ACAD. SCI. 1176–81 (2008)
(“Korlach”). Korlach was cited by Appellant as evidence that “polymerase
enzymes exhibit extensive nonspecific binding to metals, even after washing
of the metal.” App. Br. 10.
6 Declaration of Dr. Natasha Popovich under 37 C.F.R. § 1.132, signed May
2, 2016 (“Popovich Decl.”).
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despite the high concentration of fluorescently labeled
nucleotides present in solution.

App. Br. 6. Nor, Appellant asserts, is binding enzyme to aluminum
inconsistent with Figure 1 of Levene because Figure 1 is just “a schematic,
intended to illustrate the principle of operation of a ZMW.” App. Br. 7.
As stated in In re Oetiker, 977 F.2d 1443, 1445 (Fed. Cir. 1992):
“[T]he examiner bears the initial burden . . . of presenting a prima facie case
of unpatentability.” Appellant has persuaded us that the Examiner has not
carried the burden of establishing that the claimed invention was anticipated
by Levene.
We begin by construing the phrase “substantially free of the enzyme.”
The Specification states:
In the context of the foregoing, molecules of interest may be
described as being preferentially located in a particular region,
or localized substantially in a given region. It will be
appreciated that use of the term preferentially is meant to
indicate that the molecule is localized in a given location at a
concentration or surface density that exceeds that of other
locations in which it is not preferentially localized. Thus
preferential immobilization of a given molecule in a first region
will mean that the molecule is present in such region at a higher
density or concentration than in other regions. Density in such
regions may be as much as 20% greater, 30% greater, 50%
greater, 100% greater, or upwards of 200%, up to 1000% or
more of the concentration or density in other regions, and in
some cases 100 times greater, 1000 times greater or more.
Similar meaning is generally applicable to indications that a
given molecule is substantially only located in a given region.
Spec. ¶ 58. Accordingly, we understand the phrase “substantially free of the
enzyme,” as used in the context of claim 1, to mean that the enzyme bound
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to the bottom surface of the claimed well is present at a higher density or
concentration than in other regions.
Having construed the claims, we next turn to the disclosure of the
asserted art. Levene’s discloses ZMW “manufactured as small holes in an
89-nm thick film of aluminum on fused silica coverslips.” Levene, 684.
Levene further discloses that Sequenase, a polymerase, binds to silica
“nonspecifically.” Id.; see Levene, Supplementary Materials, http://science.
sciencemag.org/content/sci/suppl/2003/01/29/299.5607.682.DC1/Levene.SO
M.pdf, (2003). Korlach establishes that, under circumstances like those
described in Levene, phi29 polymerase may remain bound to aluminum,
even after the aluminum is washed with a buffer. Korlach 1178–79. In the
absence of evidence to the contrary, these teachings suggest that another
species of polymerase – the Sequenase disclosed in Levene – will also bind
to aluminum. While the Examiner discounts the evidence provided by
Korlach on the ground that Korlach used a different polymerase enzyme and
a different washing buffer, it is the Examiner’s burden to establish a prima
facie case of unpatentability. Under the Examiner’s anticipation rationale,
this requires establishing that portions of Levene’s ZMW arrays other than
the bottom surface of the wells were substantially free of polymerase bound
to the aluminum component of Levene’s substrate. Based on the record
before us, the Examiner has not established this rationale.
Figure 1 of Levene, relied upon by the Examiner as not showing
enzyme bound to aluminum, does little to support the Examiner’s
anticipation rationale because, as Appellant points out, Figure 1 is only a
schematic and does not speak to the presence of enzyme outside of the single
ZMW depicted. Levene’s teaching that unbound enzyme is washed away is
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similarly not persuasive when considered together with Korlach’s example
of a polymerase binding to aluminum.
Accordingly, we reverse the Examiner’s rejection of claims 1–4 and
6–9 as anticipated by Levene.
OBVIOUSNESS
Both of the Examiner’s obviousness rejections apply Levene as
discussed above with respect to the Examiner’s anticipation rejection. The
Examiner does not rely upon the additional art cited in the two obviousness
rejections to remedy the deficiency discussed above. Accordingly, we
reverse the Examiner’s rejection of claims 1–9 and 11 as obvious over the
combination of Levene and Hofer and the Examiner’s rejection of claims 1–
13 as obvious over the combination of Levene, Hofer, and Sluka.
DOUBLE PATENTING
The Examiner’s double patenting rejection applies Levene as
discussed above with respect to the Examiner’s anticipation rejection. The
Examiner does not rely upon the ’146 patent to remedy the deficiency
discussed above. Accordingly, we reverse the Examiner’s rejection of
claims 1, 6, and 7 on the ground of non-statutory obviousness-type double
patenting over the ’146 patent in view of Levene.

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SUMMARY
In summary:
Claims
Rejected
35 U.S.C. § Basis Affirmed Reversed
1–4, 6–9 102(b) Levene

1–4, 6–9
1–9, 11 103(a) Levene and
Hofer
1–9, 11
1–13 103(a) 103(a)
Levene,
Hofer, and
Sluka
1–13
1, 6, 7 obviousness-
type double
patenting
1, 6, and 7
Overall
Outcome
1–13

REVERSED