CORNEAL PRESERVATION STORAGE DR REKHA Lions Eye hospital

CORNEAL PRESERVATION & STORAGE DR. REKHA. Lions Eye hospital BANGALORE

IMPORTANCE OF STORAGE SUCCESS OF PENENTRATING KERATOPLASTY DEPEND UPON CORNEAL ENDOTHELIAL INTEGRITY.

CORNEAL STORAGE SYSTEM AIM w To maintain the endothelial viability & integrity. w Prevent corneal autolysis during storage w Long term corneal storage.

HISTORY w First corneal preservation. w 1911 Mgitot in J. A. M. A. w Stored cornea in Hemolyzed blood serum at 5 -7 deg. centigrade. w Cornea viable for 2 days

STORAGE TYPES SHORT-4 deg. C(days) w Moist chamber(1 day), M. K. Media(2 -4 days) w w INTERMEDIATE-14 days- 4 deg. C(wks) K-Sol, Dexsol, Optisol. GS, Procell, Eusol-C LONG(months) European. Organ culture-120 days(37 deg. C) Eurosol-31 deg. C-28 days(New) Cryoprev. -Unlimited-(-80 deg. C)

MOIST CHAMBER METHOD w w w w Filatov &Castroviejo. Whole globe is stored in a sterile jar. Jar has saline Humidification. Temp of 4 degrees C. Popular untill 1970. Simple & in- expensive. Grafting on emergency basis.

Mc. CAREY&KUFMAN STORAGE Corneo-scleral excision is necessary& immersed with endothelial side up. w Contains Tc 199, dextran-40 1%, PH 7. 0 -7. 5, Osmolality 295=355 & gentamycin sulphate 75 -150 micro gm/ml , HEPES as buffer. Phenol red as indicator. w Scheduling of surgery

STORAGE MEDIA K-SOL: Tc 199, MEM&Earles media, HEPES, Gentamicin, Chondroitin sulphate 2. 5%. 1988 proprionibacteria contamination DEXOL: MEM, 1. 35%Chondrotin Sulphate, 1 m. M Sodium pyruvate, 1 m. M non- essential aminoacids, Antioxidants, 1%dextran 40.

OPTISOL. G. S. w w w Introduced in 1991 Contents MEM, 1. 35%Chondrotin Sulphate, 1 m. MSodiumpyruvate, 1 m. M non-essential aminoacids, Antioxidants, 1%dextran 40, ATP, Iron, cholestrol, Lhydroxyproline, Vitamine, 2 antibiotics. Genticyn, Stepomycin. Expensive

EUSOL-C Store at 4 deg. C w Storage time 14 days, SIMILAR TO DEXOL w CONTENTS w Dextran, Sodium Piruvate, Glucose, Essential &non-essential aminoacids, mineral salts, Vitamins, Gentamin, hepesbuffer, Bicarbonate, Phenol Red. w w

FUNCTIONS OF CONTENTS BASE MEDIA a)Tissue-culture 199 aminoacids, salts, buffer&energy to support life. b)Minimum Essential Media(MEM) with Earles salt- matches natural salts &buffers of human tissue, electrolites similar to aqueous

FUNCTIONS OF CONTENTS BUFFER HEPES(N-2 -hydroxyethyl-piperazine-N 2 ethane-sulfonic acid) -used for cultivation of sensitive mammalian cells -provides optimal physiological Ph in the range 7 -7. 2

COMMON FEATURES OF MEDIA ANTIBIOTIC Gentamicin only /&Streptomycin DEXTRAN Polysaccharide, negatively charged. Prevents tissue swelling. Used alone does not provide sufficient tissue stability & viability for extended storage.

COMMON FEATURES OF MEDIA CHONDROTIN SULFATE Polysaccharide, negatively charged Endothelial integrity&acts as a osmotic agent. Superior to dextran. Longer storage time ENRICHED Various salts, aminoacids essential &nonessential, vit. , phosphates &antioxidents to enhance cell & tissue viability & health during storage.

COMMON MEDIA FEATURES ATP precursors--> Provide energy for pumping function. Anti-Oxidents--->Neutralize metabolic waste, maintain DNS synthesis Vitamins-->Provides additional nutritional cell supplements. Sodium Piruvate&Glucose--> Energy supply Colour Phenol Red-->Visual aid for p. H indication.

CHECK MEDIA The cornea which is stored in MKmedia, Dexol, optisol. GS. Thefollowing parameters to be checked before using Intact seal, Expiry date, Turbity, Colour (rose red) , precipitates & FB. COLOUR CHANGE INDICATES Yellow-- Bacterial contamination. Red--- Unacceptable Ph. Cloudy-- Contamination

Ph COLOUR REFERENCE CHART 7. 00 w 7. 25 7. 50 contimanation

PROCELL MEM, 1. 35%chondrotin sulphate, 1 m. M sodium pyruvate 1 m. M, Non-essential aminoacids, Antioxidants, Dextran 40, Humen insulin 10 mic. g/ml & Human epidermal growth factor(h. EGF 10 ng/ml) to improve long term endothelial survival after PKP. (experiments have not proved it, opth 1994 feb 101(2)252 -9 Lass. JH et all)

ORGAN CULTURE w w 1936, by Archer & Trevor-Roper. Being used since 1974. Refrigeration not required. Complicated, expensive & well trained microbiologist. EUROPEAN ORGAN CULTURE 31 -37 deg. C, EMEM, IMDM , HEPES, Na. HCO 3, Peni, Strep, Ampo-B, BOVINE SERUM 2 -10 ML, Dextran 4 -10%

ORGAN CULTURE contents Eagles media, Earles salt with out Lglutamine, L-glutamine 1%final conc. , Decomlemented calf serum 10%final conc. Penicilllin 100 units/ml, , Gentamicin 100 microgram/ml. Amphoterician. B 0. 25% microgram/ml.

TISSUE CULTURE MEDIA EUROSOL New Duration of storage 28 days. Temp 31 deg. C. Better then the European culture media. Maintains intact endothelium, epithelium & Keratocyte.

ORGAN CULTURE ADVANTAGES: Endothelial integrity 120 days, Epithelial integrity 35 days, Culture report prior to transplantation, stored at 4 deg, centi. DISADVANTAGES: Cornea thickens with time, Microbial contamination, Complicated expensive techniques, Greater loss of endothelial after. PK.

CRYOPRESERVATION Kaufman &Capella. 1954 w Cornea is stored in Liquid Nitrogen with Dimethyl. Sulfoxide(DMSO). w DMSO, Prevents intracellular damage by ice crystals. w Not a procedure used. Holds a lot of research interest. w

CRYOPRESERVATION ADVANTAGES: Long-term storage at 80 deg. C. DISADVANTAGES: Technically difficult, Limited tissue tolerance to freezthaw. Rapid thaw at 37 deg. C. w 1964 w 18 months w Tissue strored in Liquid nitrogen at --197 deg C w At the time of thawing the cial containing cornea is placed in water bath at temp of 60 dec C at 1 min. w Trained techiques for Freezing & thawing

GLYCERINE PRESERVATION Patch graft or Lamellar Keraoplasty. Corneo-scleral button. 100%glycerine. Enothelium is nonviable Use with in 1 year. Stored at room temp.

NOTE w HYPOTHERMIC (CS+Hyperkalaemic) w Storage 2 wks, epithelial not well preserved, not used w ORGAN CULTURE w w At one time predominating in Europe Organ culture storage-4 wks w w CRYOPRESERVATION Not used, tissure strored for emergency use

FUTURE w w w w Improve endothelial cell layer viability &compact stroma. Maintain Keratocyte viability. Reduce corneal autolysis during storage. Thin cornea in spite of long time storage. No contamenation Easy to use & transport Inexpensive.

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