An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats

Purpose Since adult rats are used in pre-clinical studies, and due to the necessity of investigating the side-effects of drugs on RPE cells in vitro, there is a great need for primary RPE cells from these animals. The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. No differences were observed between albino and pigmented RPE cells. Ausführliche Beschreibung

Gespeichert in:

Autor*in:	Langenfeld, Analena [verfasserIn] Julien, Sylvie [verfasserIn] Schraermeyer, Ulrich [verfasserIn]

Format:	E-Artikel
Sprache:	Englisch

Erschienen:	2015

Schlagwörter:	RPE isolation Retinal adhesion Rat Novel technique Immunocytochemistry

Übergeordnetes Werk:	Enthalten in: Graefe's archive for clinical and experimental ophthalmology - Berlin : Springer, 1854, 253(2015), 9 vom: 26. Apr., Seite 1493-1502
Übergeordnetes Werk:	volume:253 ; year:2015 ; number:9 ; day:26 ; month:04 ; pages:1493-1502

Links:	Volltext

DOI / URN:	10.1007/s00417-015-3011-5

Katalog-ID:	SPR005428130

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245	1	3	\|a An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats
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520			\|a Purpose Since adult rats are used in pre-clinical studies, and due to the necessity of investigating the side-effects of drugs on RPE cells in vitro, there is a great need for primary RPE cells from these animals. The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. No differences were observed between albino and pigmented RPE cells.
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700	1		\|a Julien, Sylvie \|e verfasserin \|4 aut
700	1		\|a Schraermeyer, Ulrich \|e verfasserin \|4 aut
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author_variant	a l al s j sj u s us
matchkey_str	article:1435702X:2015----::nmrvdehdotesltoadutrortnlimneih
hierarchy_sort_str	2015
bklnumber	44.95
publishDate	2015
allfields	10.1007/s00417-015-3011-5 doi (DE-627)SPR005428130 (SPR)s00417-015-3011-5-e DE-627 ger DE-627 rakwb eng 610 ASE 44.95 bkl Langenfeld, Analena verfasserin aut An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats 2015 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Purpose Since adult rats are used in pre-clinical studies, and due to the necessity of investigating the side-effects of drugs on RPE cells in vitro, there is a great need for primary RPE cells from these animals. The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. No differences were observed between albino and pigmented RPE cells. RPE isolation (dpeaa)DE-He213 Retinal adhesion (dpeaa)DE-He213 Rat (dpeaa)DE-He213 Novel technique (dpeaa)DE-He213 Immunocytochemistry (dpeaa)DE-He213 Julien, Sylvie verfasserin aut Schraermeyer, Ulrich verfasserin aut Enthalten in Graefe's archive for clinical and experimental ophthalmology Berlin : Springer, 1854 253(2015), 9 vom: 26. Apr., Seite 1493-1502 (DE-627)253723728 (DE-600)1459159-5 1435-702X nnns volume:253 year:2015 number:9 day:26 month:04 pages:1493-1502 https://dx.doi.org/10.1007/s00417-015-3011-5 lizenzpflichtig Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_32 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_90 GBV_ILN_95 GBV_ILN_100 GBV_ILN_101 GBV_ILN_105 GBV_ILN_110 GBV_ILN_120 GBV_ILN_138 GBV_ILN_150 GBV_ILN_151 GBV_ILN_152 GBV_ILN_161 GBV_ILN_170 GBV_ILN_171 GBV_ILN_187 GBV_ILN_213 GBV_ILN_224 GBV_ILN_230 GBV_ILN_250 GBV_ILN_267 GBV_ILN_281 GBV_ILN_285 GBV_ILN_293 GBV_ILN_370 GBV_ILN_602 GBV_ILN_636 GBV_ILN_702 GBV_ILN_711 GBV_ILN_2001 GBV_ILN_2003 GBV_ILN_2004 GBV_ILN_2005 GBV_ILN_2006 GBV_ILN_2007 GBV_ILN_2008 GBV_ILN_2009 GBV_ILN_2010 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2015 GBV_ILN_2020 GBV_ILN_2021 GBV_ILN_2025 GBV_ILN_2026 GBV_ILN_2027 GBV_ILN_2031 GBV_ILN_2034 GBV_ILN_2037 GBV_ILN_2038 GBV_ILN_2039 GBV_ILN_2044 GBV_ILN_2048 GBV_ILN_2049 GBV_ILN_2050 GBV_ILN_2055 GBV_ILN_2057 GBV_ILN_2059 GBV_ILN_2061 GBV_ILN_2064 GBV_ILN_2065 GBV_ILN_2068 GBV_ILN_2070 GBV_ILN_2086 GBV_ILN_2088 GBV_ILN_2093 GBV_ILN_2106 GBV_ILN_2107 GBV_ILN_2108 GBV_ILN_2110 GBV_ILN_2111 GBV_ILN_2112 GBV_ILN_2113 GBV_ILN_2116 GBV_ILN_2118 GBV_ILN_2119 GBV_ILN_2122 GBV_ILN_2129 GBV_ILN_2143 GBV_ILN_2144 GBV_ILN_2147 GBV_ILN_2148 GBV_ILN_2152 GBV_ILN_2153 GBV_ILN_2188 GBV_ILN_2190 GBV_ILN_2232 GBV_ILN_2336 GBV_ILN_2446 GBV_ILN_2470 GBV_ILN_2472 GBV_ILN_2507 GBV_ILN_2522 GBV_ILN_2548 GBV_ILN_4012 GBV_ILN_4035 GBV_ILN_4037 GBV_ILN_4046 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4242 GBV_ILN_4246 GBV_ILN_4249 GBV_ILN_4251 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4326 GBV_ILN_4328 GBV_ILN_4333 GBV_ILN_4334 GBV_ILN_4335 GBV_ILN_4336 GBV_ILN_4338 GBV_ILN_4393 GBV_ILN_4700 44.95 ASE AR 253 2015 9 26 04 1493-1502
spelling	10.1007/s00417-015-3011-5 doi (DE-627)SPR005428130 (SPR)s00417-015-3011-5-e DE-627 ger DE-627 rakwb eng 610 ASE 44.95 bkl Langenfeld, Analena verfasserin aut An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats 2015 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Purpose Since adult rats are used in pre-clinical studies, and due to the necessity of investigating the side-effects of drugs on RPE cells in vitro, there is a great need for primary RPE cells from these animals. The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. No differences were observed between albino and pigmented RPE cells. RPE isolation (dpeaa)DE-He213 Retinal adhesion (dpeaa)DE-He213 Rat (dpeaa)DE-He213 Novel technique (dpeaa)DE-He213 Immunocytochemistry (dpeaa)DE-He213 Julien, Sylvie verfasserin aut Schraermeyer, Ulrich verfasserin aut Enthalten in Graefe's archive for clinical and experimental ophthalmology Berlin : Springer, 1854 253(2015), 9 vom: 26. Apr., Seite 1493-1502 (DE-627)253723728 (DE-600)1459159-5 1435-702X nnns volume:253 year:2015 number:9 day:26 month:04 pages:1493-1502 https://dx.doi.org/10.1007/s00417-015-3011-5 lizenzpflichtig Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_32 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_90 GBV_ILN_95 GBV_ILN_100 GBV_ILN_101 GBV_ILN_105 GBV_ILN_110 GBV_ILN_120 GBV_ILN_138 GBV_ILN_150 GBV_ILN_151 GBV_ILN_152 GBV_ILN_161 GBV_ILN_170 GBV_ILN_171 GBV_ILN_187 GBV_ILN_213 GBV_ILN_224 GBV_ILN_230 GBV_ILN_250 GBV_ILN_267 GBV_ILN_281 GBV_ILN_285 GBV_ILN_293 GBV_ILN_370 GBV_ILN_602 GBV_ILN_636 GBV_ILN_702 GBV_ILN_711 GBV_ILN_2001 GBV_ILN_2003 GBV_ILN_2004 GBV_ILN_2005 GBV_ILN_2006 GBV_ILN_2007 GBV_ILN_2008 GBV_ILN_2009 GBV_ILN_2010 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2015 GBV_ILN_2020 GBV_ILN_2021 GBV_ILN_2025 GBV_ILN_2026 GBV_ILN_2027 GBV_ILN_2031 GBV_ILN_2034 GBV_ILN_2037 GBV_ILN_2038 GBV_ILN_2039 GBV_ILN_2044 GBV_ILN_2048 GBV_ILN_2049 GBV_ILN_2050 GBV_ILN_2055 GBV_ILN_2057 GBV_ILN_2059 GBV_ILN_2061 GBV_ILN_2064 GBV_ILN_2065 GBV_ILN_2068 GBV_ILN_2070 GBV_ILN_2086 GBV_ILN_2088 GBV_ILN_2093 GBV_ILN_2106 GBV_ILN_2107 GBV_ILN_2108 GBV_ILN_2110 GBV_ILN_2111 GBV_ILN_2112 GBV_ILN_2113 GBV_ILN_2116 GBV_ILN_2118 GBV_ILN_2119 GBV_ILN_2122 GBV_ILN_2129 GBV_ILN_2143 GBV_ILN_2144 GBV_ILN_2147 GBV_ILN_2148 GBV_ILN_2152 GBV_ILN_2153 GBV_ILN_2188 GBV_ILN_2190 GBV_ILN_2232 GBV_ILN_2336 GBV_ILN_2446 GBV_ILN_2470 GBV_ILN_2472 GBV_ILN_2507 GBV_ILN_2522 GBV_ILN_2548 GBV_ILN_4012 GBV_ILN_4035 GBV_ILN_4037 GBV_ILN_4046 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4242 GBV_ILN_4246 GBV_ILN_4249 GBV_ILN_4251 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4326 GBV_ILN_4328 GBV_ILN_4333 GBV_ILN_4334 GBV_ILN_4335 GBV_ILN_4336 GBV_ILN_4338 GBV_ILN_4393 GBV_ILN_4700 44.95 ASE AR 253 2015 9 26 04 1493-1502
allfields_unstemmed	10.1007/s00417-015-3011-5 doi (DE-627)SPR005428130 (SPR)s00417-015-3011-5-e DE-627 ger DE-627 rakwb eng 610 ASE 44.95 bkl Langenfeld, Analena verfasserin aut An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats 2015 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Purpose Since adult rats are used in pre-clinical studies, and due to the necessity of investigating the side-effects of drugs on RPE cells in vitro, there is a great need for primary RPE cells from these animals. The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. No differences were observed between albino and pigmented RPE cells. RPE isolation (dpeaa)DE-He213 Retinal adhesion (dpeaa)DE-He213 Rat (dpeaa)DE-He213 Novel technique (dpeaa)DE-He213 Immunocytochemistry (dpeaa)DE-He213 Julien, Sylvie verfasserin aut Schraermeyer, Ulrich verfasserin aut Enthalten in Graefe's archive for clinical and experimental ophthalmology Berlin : Springer, 1854 253(2015), 9 vom: 26. Apr., Seite 1493-1502 (DE-627)253723728 (DE-600)1459159-5 1435-702X nnns volume:253 year:2015 number:9 day:26 month:04 pages:1493-1502 https://dx.doi.org/10.1007/s00417-015-3011-5 lizenzpflichtig Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_32 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_90 GBV_ILN_95 GBV_ILN_100 GBV_ILN_101 GBV_ILN_105 GBV_ILN_110 GBV_ILN_120 GBV_ILN_138 GBV_ILN_150 GBV_ILN_151 GBV_ILN_152 GBV_ILN_161 GBV_ILN_170 GBV_ILN_171 GBV_ILN_187 GBV_ILN_213 GBV_ILN_224 GBV_ILN_230 GBV_ILN_250 GBV_ILN_267 GBV_ILN_281 GBV_ILN_285 GBV_ILN_293 GBV_ILN_370 GBV_ILN_602 GBV_ILN_636 GBV_ILN_702 GBV_ILN_711 GBV_ILN_2001 GBV_ILN_2003 GBV_ILN_2004 GBV_ILN_2005 GBV_ILN_2006 GBV_ILN_2007 GBV_ILN_2008 GBV_ILN_2009 GBV_ILN_2010 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2015 GBV_ILN_2020 GBV_ILN_2021 GBV_ILN_2025 GBV_ILN_2026 GBV_ILN_2027 GBV_ILN_2031 GBV_ILN_2034 GBV_ILN_2037 GBV_ILN_2038 GBV_ILN_2039 GBV_ILN_2044 GBV_ILN_2048 GBV_ILN_2049 GBV_ILN_2050 GBV_ILN_2055 GBV_ILN_2057 GBV_ILN_2059 GBV_ILN_2061 GBV_ILN_2064 GBV_ILN_2065 GBV_ILN_2068 GBV_ILN_2070 GBV_ILN_2086 GBV_ILN_2088 GBV_ILN_2093 GBV_ILN_2106 GBV_ILN_2107 GBV_ILN_2108 GBV_ILN_2110 GBV_ILN_2111 GBV_ILN_2112 GBV_ILN_2113 GBV_ILN_2116 GBV_ILN_2118 GBV_ILN_2119 GBV_ILN_2122 GBV_ILN_2129 GBV_ILN_2143 GBV_ILN_2144 GBV_ILN_2147 GBV_ILN_2148 GBV_ILN_2152 GBV_ILN_2153 GBV_ILN_2188 GBV_ILN_2190 GBV_ILN_2232 GBV_ILN_2336 GBV_ILN_2446 GBV_ILN_2470 GBV_ILN_2472 GBV_ILN_2507 GBV_ILN_2522 GBV_ILN_2548 GBV_ILN_4012 GBV_ILN_4035 GBV_ILN_4037 GBV_ILN_4046 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4242 GBV_ILN_4246 GBV_ILN_4249 GBV_ILN_4251 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4326 GBV_ILN_4328 GBV_ILN_4333 GBV_ILN_4334 GBV_ILN_4335 GBV_ILN_4336 GBV_ILN_4338 GBV_ILN_4393 GBV_ILN_4700 44.95 ASE AR 253 2015 9 26 04 1493-1502
allfieldsGer	10.1007/s00417-015-3011-5 doi (DE-627)SPR005428130 (SPR)s00417-015-3011-5-e DE-627 ger DE-627 rakwb eng 610 ASE 44.95 bkl Langenfeld, Analena verfasserin aut An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats 2015 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Purpose Since adult rats are used in pre-clinical studies, and due to the necessity of investigating the side-effects of drugs on RPE cells in vitro, there is a great need for primary RPE cells from these animals. The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. No differences were observed between albino and pigmented RPE cells. RPE isolation (dpeaa)DE-He213 Retinal adhesion (dpeaa)DE-He213 Rat (dpeaa)DE-He213 Novel technique (dpeaa)DE-He213 Immunocytochemistry (dpeaa)DE-He213 Julien, Sylvie verfasserin aut Schraermeyer, Ulrich verfasserin aut Enthalten in Graefe's archive for clinical and experimental ophthalmology Berlin : Springer, 1854 253(2015), 9 vom: 26. Apr., Seite 1493-1502 (DE-627)253723728 (DE-600)1459159-5 1435-702X nnns volume:253 year:2015 number:9 day:26 month:04 pages:1493-1502 https://dx.doi.org/10.1007/s00417-015-3011-5 lizenzpflichtig Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_32 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_90 GBV_ILN_95 GBV_ILN_100 GBV_ILN_101 GBV_ILN_105 GBV_ILN_110 GBV_ILN_120 GBV_ILN_138 GBV_ILN_150 GBV_ILN_151 GBV_ILN_152 GBV_ILN_161 GBV_ILN_170 GBV_ILN_171 GBV_ILN_187 GBV_ILN_213 GBV_ILN_224 GBV_ILN_230 GBV_ILN_250 GBV_ILN_267 GBV_ILN_281 GBV_ILN_285 GBV_ILN_293 GBV_ILN_370 GBV_ILN_602 GBV_ILN_636 GBV_ILN_702 GBV_ILN_711 GBV_ILN_2001 GBV_ILN_2003 GBV_ILN_2004 GBV_ILN_2005 GBV_ILN_2006 GBV_ILN_2007 GBV_ILN_2008 GBV_ILN_2009 GBV_ILN_2010 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2015 GBV_ILN_2020 GBV_ILN_2021 GBV_ILN_2025 GBV_ILN_2026 GBV_ILN_2027 GBV_ILN_2031 GBV_ILN_2034 GBV_ILN_2037 GBV_ILN_2038 GBV_ILN_2039 GBV_ILN_2044 GBV_ILN_2048 GBV_ILN_2049 GBV_ILN_2050 GBV_ILN_2055 GBV_ILN_2057 GBV_ILN_2059 GBV_ILN_2061 GBV_ILN_2064 GBV_ILN_2065 GBV_ILN_2068 GBV_ILN_2070 GBV_ILN_2086 GBV_ILN_2088 GBV_ILN_2093 GBV_ILN_2106 GBV_ILN_2107 GBV_ILN_2108 GBV_ILN_2110 GBV_ILN_2111 GBV_ILN_2112 GBV_ILN_2113 GBV_ILN_2116 GBV_ILN_2118 GBV_ILN_2119 GBV_ILN_2122 GBV_ILN_2129 GBV_ILN_2143 GBV_ILN_2144 GBV_ILN_2147 GBV_ILN_2148 GBV_ILN_2152 GBV_ILN_2153 GBV_ILN_2188 GBV_ILN_2190 GBV_ILN_2232 GBV_ILN_2336 GBV_ILN_2446 GBV_ILN_2470 GBV_ILN_2472 GBV_ILN_2507 GBV_ILN_2522 GBV_ILN_2548 GBV_ILN_4012 GBV_ILN_4035 GBV_ILN_4037 GBV_ILN_4046 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4242 GBV_ILN_4246 GBV_ILN_4249 GBV_ILN_4251 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4326 GBV_ILN_4328 GBV_ILN_4333 GBV_ILN_4334 GBV_ILN_4335 GBV_ILN_4336 GBV_ILN_4338 GBV_ILN_4393 GBV_ILN_4700 44.95 ASE AR 253 2015 9 26 04 1493-1502
allfieldsSound	10.1007/s00417-015-3011-5 doi (DE-627)SPR005428130 (SPR)s00417-015-3011-5-e DE-627 ger DE-627 rakwb eng 610 ASE 44.95 bkl Langenfeld, Analena verfasserin aut An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats 2015 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Purpose Since adult rats are used in pre-clinical studies, and due to the necessity of investigating the side-effects of drugs on RPE cells in vitro, there is a great need for primary RPE cells from these animals. The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. No differences were observed between albino and pigmented RPE cells. RPE isolation (dpeaa)DE-He213 Retinal adhesion (dpeaa)DE-He213 Rat (dpeaa)DE-He213 Novel technique (dpeaa)DE-He213 Immunocytochemistry (dpeaa)DE-He213 Julien, Sylvie verfasserin aut Schraermeyer, Ulrich verfasserin aut Enthalten in Graefe's archive for clinical and experimental ophthalmology Berlin : Springer, 1854 253(2015), 9 vom: 26. Apr., Seite 1493-1502 (DE-627)253723728 (DE-600)1459159-5 1435-702X nnns volume:253 year:2015 number:9 day:26 month:04 pages:1493-1502 https://dx.doi.org/10.1007/s00417-015-3011-5 lizenzpflichtig Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_32 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_90 GBV_ILN_95 GBV_ILN_100 GBV_ILN_101 GBV_ILN_105 GBV_ILN_110 GBV_ILN_120 GBV_ILN_138 GBV_ILN_150 GBV_ILN_151 GBV_ILN_152 GBV_ILN_161 GBV_ILN_170 GBV_ILN_171 GBV_ILN_187 GBV_ILN_213 GBV_ILN_224 GBV_ILN_230 GBV_ILN_250 GBV_ILN_267 GBV_ILN_281 GBV_ILN_285 GBV_ILN_293 GBV_ILN_370 GBV_ILN_602 GBV_ILN_636 GBV_ILN_702 GBV_ILN_711 GBV_ILN_2001 GBV_ILN_2003 GBV_ILN_2004 GBV_ILN_2005 GBV_ILN_2006 GBV_ILN_2007 GBV_ILN_2008 GBV_ILN_2009 GBV_ILN_2010 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2015 GBV_ILN_2020 GBV_ILN_2021 GBV_ILN_2025 GBV_ILN_2026 GBV_ILN_2027 GBV_ILN_2031 GBV_ILN_2034 GBV_ILN_2037 GBV_ILN_2038 GBV_ILN_2039 GBV_ILN_2044 GBV_ILN_2048 GBV_ILN_2049 GBV_ILN_2050 GBV_ILN_2055 GBV_ILN_2057 GBV_ILN_2059 GBV_ILN_2061 GBV_ILN_2064 GBV_ILN_2065 GBV_ILN_2068 GBV_ILN_2070 GBV_ILN_2086 GBV_ILN_2088 GBV_ILN_2093 GBV_ILN_2106 GBV_ILN_2107 GBV_ILN_2108 GBV_ILN_2110 GBV_ILN_2111 GBV_ILN_2112 GBV_ILN_2113 GBV_ILN_2116 GBV_ILN_2118 GBV_ILN_2119 GBV_ILN_2122 GBV_ILN_2129 GBV_ILN_2143 GBV_ILN_2144 GBV_ILN_2147 GBV_ILN_2148 GBV_ILN_2152 GBV_ILN_2153 GBV_ILN_2188 GBV_ILN_2190 GBV_ILN_2232 GBV_ILN_2336 GBV_ILN_2446 GBV_ILN_2470 GBV_ILN_2472 GBV_ILN_2507 GBV_ILN_2522 GBV_ILN_2548 GBV_ILN_4012 GBV_ILN_4035 GBV_ILN_4037 GBV_ILN_4046 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4242 GBV_ILN_4246 GBV_ILN_4249 GBV_ILN_4251 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4326 GBV_ILN_4328 GBV_ILN_4333 GBV_ILN_4334 GBV_ILN_4335 GBV_ILN_4336 GBV_ILN_4338 GBV_ILN_4393 GBV_ILN_4700 44.95 ASE AR 253 2015 9 26 04 1493-1502
language	English
source	Enthalten in Graefe's archive for clinical and experimental ophthalmology 253(2015), 9 vom: 26. Apr., Seite 1493-1502 volume:253 year:2015 number:9 day:26 month:04 pages:1493-1502
sourceStr	Enthalten in Graefe's archive for clinical and experimental ophthalmology 253(2015), 9 vom: 26. Apr., Seite 1493-1502 volume:253 year:2015 number:9 day:26 month:04 pages:1493-1502
format_phy_str_mv	Article
institution	findex.gbv.de
topic_facet	RPE isolation Retinal adhesion Rat Novel technique Immunocytochemistry
dewey-raw	610
isfreeaccess_bool	false
container_title	Graefe's archive for clinical and experimental ophthalmology
authorswithroles_txt_mv	Langenfeld, Analena @@aut@@ Julien, Sylvie @@aut@@ Schraermeyer, Ulrich @@aut@@
publishDateDaySort_date	2015-04-26T00:00:00Z
hierarchy_top_id	253723728
dewey-sort	3610
id	SPR005428130
language_de	englisch
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The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. 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author	Langenfeld, Analena
spellingShingle	Langenfeld, Analena ddc 610 bkl 44.95 misc RPE isolation misc Retinal adhesion misc Rat misc Novel technique misc Immunocytochemistry An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats
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topic_title	610 ASE 44.95 bkl An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats RPE isolation (dpeaa)DE-He213 Retinal adhesion (dpeaa)DE-He213 Rat (dpeaa)DE-He213 Novel technique (dpeaa)DE-He213 Immunocytochemistry (dpeaa)DE-He213
topic	ddc 610 bkl 44.95 misc RPE isolation misc Retinal adhesion misc Rat misc Novel technique misc Immunocytochemistry
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title	An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats
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title_full	An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats
author_sort	Langenfeld, Analena
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author_browse	Langenfeld, Analena Julien, Sylvie Schraermeyer, Ulrich
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title_sort	improved method for the isolation and culture of retinal pigment epithelial cells from adult rats
title_auth	An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats
abstract	Purpose Since adult rats are used in pre-clinical studies, and due to the necessity of investigating the side-effects of drugs on RPE cells in vitro, there is a great need for primary RPE cells from these animals. The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. No differences were observed between albino and pigmented RPE cells.
abstractGer	Purpose Since adult rats are used in pre-clinical studies, and due to the necessity of investigating the side-effects of drugs on RPE cells in vitro, there is a great need for primary RPE cells from these animals. The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. No differences were observed between albino and pigmented RPE cells.
abstract_unstemmed	Purpose Since adult rats are used in pre-clinical studies, and due to the necessity of investigating the side-effects of drugs on RPE cells in vitro, there is a great need for primary RPE cells from these animals. The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. No differences were observed between albino and pigmented RPE cells.
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container_issue	9
title_short	An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats
url	https://dx.doi.org/10.1007/s00417-015-3011-5
remote_bool	true
author2	Julien, Sylvie Schraermeyer, Ulrich
author2Str	Julien, Sylvie Schraermeyer, Ulrich
ppnlink	253723728
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isOA_txt	false
hochschulschrift_bool	false
doi_str	10.1007/s00417-015-3011-5
up_date	2024-07-03T16:15:37.844Z
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The aim of this study was to develop a reproducible and quantifiable method of isolation, culture, and maintenance of adult rat RPE cells. Moreover, potential differences between RPE cells from albino versus pigmented rats were also investigated. Methods A total of 180 pigmented rats and 340 albino rats aged 6–14 weeks were used. RPE cells were isolated and cultured for several weeks by using three different methods: 1) growing directly on flat mounts, 2) after enzymatic isolation, and 3) after they spontaneously detached from the flat mounts and continued to grow on the plastic. Yield, cell survival, and morphological characteristics were investigated using light and electron microscopy as well as immunohistochemistry. Results After 0 weeks, the yield of the first method was 30,000 cells/eye; after 2 weeks 18,000 cells/eye; and after 4 weeks 11,000 cells/eye. The yield of RPE cells was very low after enzymatic isolation in method 2 (0 weeks, 13.000 cells/eye; 2 weeks, 30,000 cells/eye; 4 weeks 38,000 cells/eye), whereas it was higher when the RPE cells spontaneously detached from the flat mounts and then continued to grow on the plastic in method three. (0 weeks, 30,000 cells/eye; 2 weeks, 314,000 cells/eye; 4 weeks, 659,000 cells/eye). The second method often showed contamination with fibroblasts, whereas the two other methods showed pure RPE cultures. The RPE cells were able to proliferate when using the second and the third method, but not when they were cultivated directly on the flat mounts (first method). Conclusion The qualitative and quantitative best method for isolating adult rat RPE cells is the culture of RPE cells which spontaneously detach from flat mounts. 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An improved method for the isolation and culture of retinal pigment epithelial cells from adult rats

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