December 2002
Volume 43, Issue 13
ARVO Annual Meeting Abstract  |   December 2002
Staining Characteristics of Preserved Human Amniotic Membrane
Author Affiliations & Notes
  • S Basti
    Department of Ophthalmology Northwestern University Medical School Chicago IL
  • DJ Hu
    Department of Ophthalmology Northwestern University Medical School Chicago IL
  • PJ Bryar
    Department of Ophthalmology Northwestern University Medical School Chicago IL
  • Footnotes
    Commercial Relationships   S. Basti, None; D.J. Hu, None; P.J. Bryar, None.
Investigative Ophthalmology & Visual Science December 2002, Vol.43, 4185. doi:
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      S Basti, DJ Hu, PJ Bryar; Staining Characteristics of Preserved Human Amniotic Membrane . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4185.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract: : Purpose: Amniotic membrane is an ultra-thin, transparent, cellophane-like membrane that is currently being used in ocular surface reconstruction. The purpose of this study is to evaluate the staining characteristics of commonly available dyes on preserved human amniotic membrane to aid in handling of amniotic membrane during transplantation. Methods: Five dyes, indocyanine green (ICG-2.5%, 1.0%, 0.5%), fluorescein (0.25%), rose bengal (1%), trypan blue (0.5%), and lissamine green B (1%) were used to stain pieces of fresh-frozen and freeze-dried amniotic membrane. The amniotic membrane was placed in 2-3 drops of dye and spread flat to ensure exposure of the entire specimen to the dye. Following staining, the membrane was washed in balanced saline solution to remove excess dye. After mounting the specimen in balanced saline, the membrane was then evaluated under magnification. Following this, the stained amniotic membranes were sectioned and examined histologically and compared with an H&E treated unstained specimen to evaluate deposition of the various stains. Membranes stained with dye were also evaluated for the persistence of staining. This was evaluated by placing stained membranes in 2.5 ml of balanced saline solution that was changed every 30 minutes for the first 6 hours. The specimens were observed every 15 minutes both grossly and under the microscope for any decrease in the degree of staining. Results: Preserved human amniotic membrane is positively stained by ICG (2.5%, 1.0%, 0.5%), 1% rose bengal, 1% lissamine green B, and 0.5% trypan blue, but not by 0.25% sodium fluorscein. Edges as well as folds of the amniotic membrane were much more easily identifiable in sections with positive staining when compared to unstained controls. Of these four dyes, only the membrane stained with 1% lissamine green B was free of stain when placed in saline for 60 minutes. ICG, trypan blue, and rose bengal continued to strongly stain the membrane even after 24 hours. Conclusion: ICG , rose bengal, lissamine green B, and trypan blue stain amniotic membrane and can aid the surgeon in handling the membrane during transplantation by clearly identifying the edges and folds in the tissue. Lissamine green B leaches out in 60 minutes, and because of this may be the dye of choice for intraoperative staining of amniotic membrane.

Keywords: 369 cornea: clinical science • 372 cornea: epithelium • 607 transplantation 

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