ORIGINAL ARTICLE

Tissue quality of eye-bankprepared precut corneas for

Descemets stripping automated endothelial keratoplasty
Brian A. Nelson, MD, Rusty J. Ritenour, MD, FRCSC
ABSTRACT RSUM
Objective: To evaluate endothelial cell density (ECD) of eye-bankprepared tissue for use in Descemets stripping automated
endothelial keratoplasty (DSAEK).
Design: Prospective case series of consecutive corneal tissue prepared for DSAEK surgery.
Participants: Sixty-seven sequential corneal-scleral tissue specimens representing 48 human donors processed for use in DSAEK
surgery by the Regional Tissue Bank (Halifax, Nova Scotia).
Methods: Corneal-scleral donor tissue was obtained by in situ recovery. ECD was recorded using the EB-3000 XYZ (HAI
Laboratories Inc, Lexington, MA) specular microscope within 24 hours of preservation. Before the tissue was dissected, the
corneal thickness was measured using the DGH-550 PACHETTE 2 (DGH Technology, Exton, PA) ultrasound pachymeter. The
dissection was performed using a 300-m Moria ALTK model microkeratome (Moria Inc). The posterior bed thickness was
measured, and the anterior flap was replaced. Endothelial cell count density was obtained after re-preservation.
Results: Complete measurements were obtained for 42 of 67 corneas. In 25 corneas it was not possible to obtain a postdissection
ECD measurement. The mean ECD before dissection was 2806 317 cells/mm
2
. The mean ECD after dissection was 2772
318 cells/mm
2
. There was an average loss of 34 cells/mm
2
(95% CI 110 to 40 cells/mm
2
, p 0.3).
Conclusions: This case series confirms that ECD is preserved when DSAEK tissue is prepared in advance of surgery by trained eye-
bank technicians in a low-volume Canadian eye bank. It was difficult to obtain clear images of the endothelial cell layer
postdissection, possibly because of tissue swelling or distortion. Sixty-six of 67 corneas included in the study were used for surgery.
Objet : valuation de la densit des cellules endothliales (DCE) des tissus prpars par la banque dyeux pour la kratoplastie
endothliale automatise avec dcapage de la Descemet (KEADD).
Nature : Srie de cas prospectifs de tissus cornens conscutifs prpars pour la chirurgie de KEADD.
Participants : 67 spcimens squentiels de tissu sclro-cornen provenant de 48 donneurs humains pour utilisation dans la
chirurgie KEADD, de la Banque rgionale de tissu de Halifax, Nouvelle-cosse.
Mthodes : Le tissu sclro-cornen des donneurs a t obtenu par rcupration sur place. La DCE a t releve avec le
microscope spculaire EB-3000 XYZ (HAI Labs, USA) sous 24 heures de prservation. Avant la dissection du tissu, lpaisseur
de la corne avait t mesure avec le pachymtre lultrason DGH-550 PACHETTE 2 (DGH Technology, USA). La dissection a
t excute avec un microkratome de 300 um de modle Moria ALTK (Moria Inc., USA). Lpaisseur du lit postrieur a t
mesure et le lambeau antrieur, replac. La DCE a t obtenu la suite de la reprservation.
Rsultats : Les mesures compltes ont t obtenues pour 42 cornes sur 67. Dans 25 cornes, il na pas t possible dobtenir
une mesure DCE aprs la dissection. Avant la dissection, la DCE moyenne tait de 2806 + 317 cellules/2mm. La moyenne de
DCE aprs dissection tait de 2772 + 318 cellules/2mm (95 % CI - 110 40 cellules/2mm, p=0,3). En moyenne, il y a eu une
perte de 34 cellules/mm2 (95% IC -110 40 cellules/mm2 p=0.3).
Conclusion : Cette srie de cas confirme que la DCE est conserve lorsque le tissu de la KEADD est prpar davance en petit
volume par des techniciens qualifis dans une banque dyeux canadienne pour la chirurgie. Il tait difficile dobtenir des images
claires de la couche cellulaire endothliale aprs la dissection, ce qui est peut-tre d lenflure ou la distorsion du tissu. 66 des
67 cornes incluses dans ltude ont servi la chirurgie.
Endothelial keratoplasty (EK) describes corneal transplan-
tation surgeries that use a posterior lamellar graft of donor
endothelium, either with Descemets membrane alone or
with stromal tissue. EK is an area of rapid development
since the rst technique was described by Melles et al.
1
in
1999. Advancements in technique have greatly improved
visual outcomes
2
and reduced graft failure rates.
3,4
EK is
seeing rapidly growing utilization rates compared with
penetrating keratoplasty for endothelial dysfunction.
5,6
For Descemets stripping automated endothelial kerato-
plasty (DSAEK), the graft dissection is most commonly
performed using a microkeratome,
7
with some centres
using a femtosecond laser.
810
Worldwide popularity of
DSAEK has surpassed that of other EK techniques because
of the simplicity of donor tissue preparation, increasing
supply of eye-bankprepared tissue, and an increasing body
of evidence supporting the safety and efcacy of the
procedure. DSAEK has been shown to reduce incidence
of donor perforation and reduce recovery time without any
reduction in visual outcome over manual dissection.
3
In
DSAEK, the surgeon either prepares the donor tissue at the
time of surgery or uses eye-bankprepared tissue.
11,12
Tissue prepared before surgery by a technician reduces
the length and complexity of DSEAK surgery. Surgeon-
prepared and eye-bankprepared tissue have been shown to
have similar nal endothelial cell loss, visual and refractive
outcomes, and dislocation rates.
11,13
Predissected tissue
carries several other advantages, including avoidance of
From the Dalhousie University, Halifax, N.S
Presented at the Canadian Ophthalmological Society Annual Meeting in
Vancouver, B.C., June 11, 2011.
Originally received Jan. 26, 2013. Final revision Sep. 12, 2013. Accepted
Sep. 17, 2013
Correspondence to Brian A. Nelson, MD, Dalhousie University, 1276
South Park Street, Halifax NS B3H 2Y9; brian.nelson@dal.ca
Can J Ophthalmol 2014;49:9295
0008-4182/14/$-see front matter & 2014 Canadian Ophthalmological
Society. Published by Elsevier Inc. All rights reserved.
http://dx.doi.org/10.1016/j.jcjo.2013.09.017
92 CAN J OPHTHALMOLVOL. 49, NO. 1, FEBRUARY 2014
surgical delays because of tissue damage at the time of
surgery. Technician-prepared tissue from high-volume eye
banks has been shown to have acceptable rates of endo-
thelial cell density (ECD) loss and consistent graft thick-
ness.
12,14
Kelliher et al.
14
reported 913 technician-prepared
corneas from a single eye bank and found an average gain
in ECD of 136 cells/mm
2
. Chen et al.
12
reported 80
corneas from 2 independent eye banks and found an
average loss in ECD of 99 cells/mm
2
. In Canada, because
of geographic constraints and a smaller population, there
are many eye banks with a lower tissue volume than those
reported in the literature. The purpose of this article is to
evaluate tissue quality parameters of ECD and micro-
keratome cut depth in a low-volume Canadian eye bank
with newly trained technicians performing the dissection.
METHODS
Data were prospectively collected for consecutive
DSAEK tissue processed by Regional Tissue Bank techni-
cians at the Queen Elizabeth II Health Sciences Centre
(Halifax, Nova Scotia). The Tissue Bank is approved by
Health Canada and is an International Associate Member
of the Eye Bank Association of America. Approval of the
study protocol was obtained from the local institutions
research ethics board. The rst tissue was processed by the
tissue bank technicians for transplantation in May 2010.
Consecutive tissue was included in the study until
February 2011. The 2 technicians performing the dis-
sections were trained by local ophthalmologists using
tissue not suitable for transplantation but for which
consent was obtained for research and education use. In
situ excisions of corneal-scleral rims were performed by
local technicians in Halifax and a satellite eye bank in St.
John, New Brunswick. Within 24 hours of preservation,
the tissue was evaluated by slit lamp and ECD measure-
ments were obtained using an EB-3000 XYZ specular
microscope (HAI Laboratories Inc, Lexington, MA).
Study corneas underwent the standard tissue bank
protocol for screening. In short, suitable tissue (donor
age o 70 years, ECD 4 2000) undergoes serology scre-
ening and a medical social interview with the donors next
of kin and family physician. Suitable tissues are then
offered to the cornea transplant service where the decision
is made regarding its use for DSAEK surgery. To be
considered for DSAEK, tissue must have ECD greater
than 2200, few or no corneal folds, clear stroma, and few
or no guttae. The tissue is predissected no more than 24
hours before the surgery. The tissue is mounted on a
Moria Articial Anterior Chamber (Moria Inc). The
anterior chamber is pressurized using a syringe with
balanced salt solution until the cornea is rm to digital
tonometry. The corneal epithelium is then removed using
a sterile cellulose sponge eye spear. The thickness of the
remaining donor tissue is then measured 3 times using a
DGH-550 PACHETTE 2 (DGH Technology, Lexing-
ton, MA) pachymeter. An alignment mark is made using a
marking pen in the periphery of the cornea. The cornea is
then cut using a Moria ALTK Microkeratome (Moria Inc,
Antony, France) with 300-m head, creating a free cap.
The cap is set aside and the thickness of the remaining
posterior lamella is measured 3 times. The surface is dried
using a cellulose surgical spear and the ap replaced.
Centration of the ap is conrmed using the peripheral
mark made earlier. The tissue is then preserved in Optisol-
GS (Bausch & Lomb, Rochester, NY). Tissue is evaluated
after dissection with slit-lamp biomicroscopy and ECD.
RESULTS
One hundred and thirty-nine corneas were prepared by
the eye bank for corneal transplantation during the study
period. Of these, 67 corneas representing 58 human
donors were selected by the transplant surgeon for DSAEK
preparation. All remaining tissues were used for other
corneal transplantation techniques. The donors consisted
of 33 males and 25 females (Table 1). The mean donor
age was 51 years, with a range of 12 to 69 years. The most
common cause of donor death was cardiovascular disease.
One cornea was lost during the study period because of
anterior chamber collapse during preparation. This
occurred because the tissue was initially excised with a
thin scleral rim, and an adequate seal to the articial
anterior chamber was not obtained. This cornea was not
included in the analysis. The remaining 66 corneas
included in the study were all used for surgery.
Tissue was preserved in Optisol GS. The mean time to
preservation from pronouncement of cardiac death by the
attending physician was 9.1 hours. Once preserved in
Optisol, tissue was then a mean of 7 days later (Table 2).
The mean ECD at the time of preservation was 2810 cells/
mm
2
. Mean ECD measured after preparation was 2770
cells/mm
2
in the 41 corneas where postcut measurements
were successfully obtained. The mean ECD loss was 35
cells/mm
2
, which was not statistically signicant. ECD
loss in each sequential cut is shown in Figure 1. In the 26
corneas where postcut measurements were not obtained,
the technician was unable to visualize the endothelial layer
using the specular microscope.
Tissue that had a high predissection ECD tended to
measure lower ECD postdissection. Likewise, low initial
ECD measurements led to higher nal ECD measurement
Table 1Donor characteristics
Donor sex 33 (57%) males, 25 (43%) females
Donor age, y Mean 51, range 1269
Cause of death, n (%)
Cardiovascular 19 (33%)
Cerebrovascular 7 (12%)
Lung cancer 4 (7%)
Trauma 4 (7%)
Tissue quality of precut corneas for DSAEKNelson and Ritenour
CAN J OPHTHALMOLVOL. 49, NO. 1, FEBRUARY 2014 93
in some corneas. Technician technique was expected to
improve with volume of cut procedures performed. To
quantify an improved outcome because of learning, we
compared ECD loss in the rst 33 corneas with the last 34
using the paired t test. No signicant difference was
detected. There was, however, an improvement in the
success rate of obtaining cell density measurement post-
dissection. In the rst 33 dissections, nal ECD measure-
ments were obtained in 17 corneas (51.5%). In the nal
34 dissections, nal ECD measurements were obtained in
25 corneas (73.5%).
In all corneas, the 300-m microkeratome head was
used. The expected cut depth assumed by other eye banks
is 370 m.
14
Of the 67 corneas included in the study, 60
had complete pachymetry measurements before and after
the cut. The average precut pachymetry measured imme-
diately after removal of the epithelium was 529 50 m.
The average cut depth was 347 53 m, calculated by
subtracting the posterior bed thickness from the predis-
section corneal thickness (Fig. 2). The 3 deepest cuts were
446, 459, and 540 m, and were associated with higher
predissection corneal thickness of 565, 549, and 663 m,
respectively.
DISCUSSION
The comparatively low-volume Canadian Eye Bank
studied demonstrated tissue quality metrics on par with
those reported in larger centres elsewhere.
14,15
As in other
studies, there were corneas where a physiologically impos-
sible gain in ECD was observed, as well as large losses in
ECD. These tended to occur in corneas that were outliers
in initial ECD testing and could be explained by regres-
sion to the mean or inaccurate initial measurements. This
same effect was observed by Kelliher et al.,
14
who reported
a mean ECD gain of 136 cells/mm
2
, with a standard
deviation of 305 cells/mm
2
in 913 corneas. Chen et al.
12
reported an average loss of 99 cells/mm
2
, with a standard
deviation of 179 and a range of 351 to 531 in 80 corneas
using the same specular microscope as our study. Rose
et al.
15
reported measurements in 6 corneas before and
after dissection, and demonstrated an average loss of 328
cells/mm
2
with a range of 163 to 1380 cells/mm
2
,
although the measurements were taken at 48 hours after
dissection and a EKA-98 Keratoanalyzer (Konan Medical
Inc, Rancho Palos Verdes, Calif.) was used. Terry et al.
16
reported clinical outcomes of 90 consecutive DSAEK
procedures and demonstrated that predissection ECD
had no effect on post-op ECD or the incidence of graft
dislocation and primary graft failure. In our study, large
outliers are best explained by measurement error. For
example, 1 cornea measured 655 cells/mm
2
higher after
dissection.
There was no signicant loss of ECD during tissue
handling throughout the study. Only 1 tissue was lost
during handling. The collapse of the articial chamber
during cutting was related to insufcient scleral rim tissue
when the cornea was initially excised from the donor. Our
results are comparable with the published rates of tissue
loss during dissection range from 1.5% in 913 corneas
14
to 2.5% in 100 corneas.
12
There were difculties obtain-
ing clear specular microscopy images after the dissections.
Tissue distortion and corneal edema caused by excess
manipulation of the corneal tissue may be implicated. This
is further suggested by improvement in the rate of
successful specular microscopy image acquisition in the
later dissections, when the technicians had more experi-
ence with the procedure. The technicians had extensive
experience with specular microscopy before the eye bank
began preparing DSAEK tissue. The lack of postdissection
Fig. 2Depth of microkeratome cut of sequential dissections.
Fig. 1Change in endothelial cell density (ECD) of sequential
dissections.
Table 2Tissue characteristics
Result Mean (range)
Time from death to preservation 9:04 h (3:19 to 15:45)
Time from preservation to dissection 7 2.5 days (313)
ECD at initial preservation, cells/mm
2
2810 320 (22003490)
ECD postdissection, cells/mm
2
2770 320 (22203390)
ECD change, cells/mm
2
35 (95% CI 110 to 40, p 0.3)
ECD, endothelial cell density.
Tissue quality of precut corneas for DSAEKNelson and Ritenour
94 CAN J OPHTHALMOLVOL. 49, NO. 1, FEBRUARY 2014
ECD measurements in 25 of 66 corneas introduces a
potential bias in the analysis of cell loss, because tissue that
was difcult to image may have experienced more cell loss.
The majority of literature on eye-bankprepared tissue
for DSAEK comes from high-volume surgical centres and
large eye banks in the United States and Europe. Our
study demonstrates that technicians in lower volume tissue
banks can perform the procedure effectively and safely. A
rigorous training program initiated by local corneal trans-
plant surgeons using nontransplantable human corneas
was effective. Cost savings are achieved by reducing
operating room time required by the surgeon. These data
should encourage low-volume eye banks to become
involved in DSAEK tissue preparation.
Disclosure: The authors have no proprietary or commercial
interest in any materials discussed in this article.
Acknowledgements: The authors acknowledge Paul Artes,
PhD (Department of Ophthalmology and Vision Sciences,
Dalhousie University) for assistance with statistical analysis,
Catherine Hackett (Regional Tissue Bank, QEII Health Sciences
Centre, Halifax, Nova Scotia), and Mary Gatien (New Bruns-
wick Eye Bank, Saint John, Nova Scotia).
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