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Deep lamellar keratoplasty with trypan blue intrastromal staining

Balestrazzi, Emilio MD*,a; Balestrazzi, Angelo MDb; Mosca, Luigi MDa; Balestrazzi, Alessandra MDc

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Journal of Cataract & Refractive Surgery: June 2002 - Volume 28 - Issue 6 - p 929-931
doi: 10.1016/S0886-3350(02)01225-7
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Deep lamellar keratoplasty is a recent surgical procedure in which the layers of the cornea except Descemet's membrane and the endothelium are replaced by donor tissue.1,2 A lamellar graft offers advantages over penetrating keratoplasty (PKP) such as avoiding most intraoperative complications and reducing the risk of allograft rejection because endothelium is not transplanted. Compared to a more anterior lamellar procedure, deep lamellar keratoplasty causes fewer irregularities and less scarring of the interface with consequent irregular astigmatism and defective vision. However, deep lamellar keratoplasty is difficult to perform because it is not possible to visualize the stromal dissection depth and it is easy to inadvertently perforate Descemet's membrane.

Many alternative methods have been advocated to dissect down to Descemet's membrane. These include intrastromal air injection,3 intrastromal balanced salt solution (BSS®) injection,4 and viscodissection after air injection into the anterior chamber.5,6

Trypan blue is a dye that is usually used in ophthalmic surgery to stain the anterior lens capsule to facilitate the creation of a continuous curvilinear capsulorhexis in the absence of a red fundus reflex.7 We present a deep lamellar keratoplasty technique in which the intrastromal bed is stained with trypan blue 0.02% solution, allowing good visualization of the dissection depth and decreasing the risk of perforation of Descemet's and endothelial layers.

Surgical Technique

The cornea is marked with a 12-dot corneal marker. An 8.0 mm central trephination with a depth corresponding to two thirds of the pachymetric corneal thickness is made with a Hanna suction trephine system.

A trypan blue 0.02% solution is prepared by adding 2.0 mL of a sterile balanced salt solution to 0.5 mL of a commercially available 0.1% solution. The trypan blue 0.02% solution is injected intrastromally in 4 quadrants of the central 8.0 mm corneal lenticule through a 30-gauge cannula (Figure 1).

Figure 1.
Figure 1.:
(Balestrazzi) Trypan blue 0.02% solution is injected intrastromally after a two-thirds corneal thickness trephination is made.

A superficial lamellar dissection, guided by the stained stromal fibers, is performed to the trephined edge with a disposable disc knife. A limbal paracentesis is created to lower intraocular pressure to prevent corneal perforation during the deep stromal dissection. An incision is made with a disc knife to the deepest stromal layers in the 4 quadrants. Each quadrant is again injected with the trypan blue solution.

The deep stromal dissection is made from the periphery to the corneal center with a smoothed spatula. The deep stromal layers are cut with a 30-degree disposable knife (Figure 2) and with corneal scissors. In the 5.0 mm central area of Descemet's membrane, the residual stromal fibers stained with trypan blue solution are removed with a disc knife.

Figure 2.
Figure 2.:
(Balestrazzi) The stromal layers are cut with a 30-degree disposable knife that is guided by the dissecting spatula to prevent perforations.

An 8.25 mm donor corneal lenticule is trephined. Its posterior surface is swabbed to remove the endothelium and Descemet's membrane. The donor lenticule is sutured into the recipient bed, first with 8 interrupted sutures and then with a double-running, 12-passage 10-0 monofilament nylon suture (Figure 3).

Figure 3.
Figure 3.:
(Balestrazzi) Final surgical outcome after the double-running, 12-passage suture is tightened.


In the past few years, surgeons have expressed great interest in deep lamellar keratoplasty as it does not lead to graft rejection. Deep lamellar keratoplasty is mainly indicated in cases of keratoconus, corneal leukomas, and corneal epithelial and stromal dystrophies. Moreover, unlike in superficial lamellar keratoplasty, the visual outcomes of deep lamellar keratoplasty are satisfactory and similar to those of PKP. Several techniques have been described to provide good visualization of Descemet's membrane, thus preventing corneal perforation and conversion to a penetrating graft.3–6

The most difficult step in deep lamellar keratoplasty is the removal of the deep stromal layers because the stromal dissection depth relative to the corneal thickness cannot be optically visualized. The injection of a trypan blue 0.02% solution into the stromal fibers enables the surgeon to visualize and remove the posterior stromal layers intraoperatively. The dye fades in the early postoperative period until it can no longer be seen (Figure 4). The results of randomized studies comparing our deep lamellar keratoplasty technique with air lamellar keratoplasty, viscolamellar keratoplasty, fluid lamellar keratoplasty, and other deep lamellar surgical techniques will determine the role and efficacy of our method.

Figure 4.
Figure 4.:
(Balestrazzi) Slitlamp image of deep lamellar keratoplasty 10 days after surgery. No residual trypan blue staining is visible.


1. Anwar M. Dissection technique in lamellar keratoplasty. Br J Ophthalmol 1972; 56:711-713
2. Barraquer JI. Lamellar keratoplasty (special techniques). Ann Ophthalmol 1972; 4:437-469
3. Price FW Jr. Air lamellar keratoplasty. Refract Corneal Surg 1989; 5:240-243
4. Amayem AF, Anwar M. Fluid lamellar keratoplasty in keratoconus. Ophthalmology 2000; 107:76-79; discussion by PR Laibson, 80
5. Melles GRJ, Lander F, Rietveld FJR, et al. A new surgical technique for deep stromal, anterior lamellar keratoplasty. Br J Ophthalmol 1999; 83:327-333
6. Melles GRJ, Remeijer L, Geerards AJM, Beekhuis WH. A quick surgical technique for deep, anterior lamellar keratoplasty using visco-dissection. Cornea 2000; 19:427-432
7. Melles GRJ, de Waard PWT, Pameyer JH, Beekhuis WH. Trypan blue capsule staining to visualize the capsulorhexis in cataract surgery. J Cataract Refract Surg 1999; 25:7-9
© 2002 by Lippincott Williams & Wilkins, Inc.