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LI Ivert, J Kong, P Gouras; Culturing Transviteally Biopsied Rabbit Retinal Pigment Epithelium . Invest. Ophthalmol. Vis. Sci. 2002;43(13):3449.
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Purpose:To determine the possibility of harvesting retinal pigment epithelium (RPE) from small transvitreal biopsies of rabbit retina, the minimum number of RPE cells needed to establish a confluent culture and the post-operative status of the biopsy site. Methods:After a local vitrectomy, a small retinal detachment is made and a smooth, flat ended glass pipette (outer /inner diameter: 0.2/0.1 mm) is introduced through the retinotomy. The pipette is connected to a foot-pedal controlled electronic micro-suction or micro-injection system to suck up RPE cells from Bruch's membrane within the detachment. Dislodged RPE cells are removed and injected into a small (7 mm diameter) culture well and subsequently monitored for RPE cell number and division. The biopsy site is followed by ophthalmoscopy, scanning laser ophthalmoscopy (SLO) and fluorescein and indocyanine green angiography. Results: Approximately 500-5000 RPE cells were removed and introduced into a culture well. Approximately 50-500 such cells (10%) attached to the culture well surface. From this population, cell division invariably occurs but only after 2-4 weeks of culturing. Only half of the cultures became confluent, invariably those with the largest number of primary cells. Angiography of the biopsy site showed window defects and choriocapillary atrophy and large choroidal vessel narrowing. The retinotomy sealed almost completely. Conclusion:It is possible to obtain sufficient RPE cells from a relatively small biopsy of rabbit RPE in vivo to establish a confluent culture of such cells. The lower the initial number of primary cells, the less is the chance that a culture becomes confluent. The RPE debridement alters both the choriocapillaris and the larger choroidal vessels.
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