Abstract
An erratum was issued for: Isolation of Primary Mouse Retinal Pigmented Epithelium Cells. The Discussion and References sections were updated.
The third paragraph of the Discussion section was updated from:
This protocol is different from other valuable and reliable published protocols for primary mouse RPE isolation13,19,20 in that it is simplified with a few steps that are easy to follow. RPE cells are able to be identified by their morphology, pigmentation, and specific RPE markers such as RPE654,5,21. Validation of the purity and contamination of Müller cells was achieved by staining isolated cells with a specific cell marker for Müller cells (vimentin)22. Many techniques have been used to evaluate the integrity of the blood-retinal barrier (BRB) in vitro, such as electron microscope (EM) examination, assessment of tight junction proteins (TJPs), FITIC dextran leakage assay, and transepithelial electrical resistance (TER) measurement that evaluates the physiological function of RPE as a monolayer8,23. RPE cells play an important role in maintaining the outer BRB, and to validate this function, we evaluated tight junction proteins (TJPs) such as Zona ocludin-1 (ZO-1) and cytoskeletal proteins like F-actin as previously published8. TJPs evaluation is a more convenient method for the evaluation of BRB integrity and barrier function which doesn't require expensive equipment that may not be available in all research labs, as opposed to EM evaluation or TER.
to:
This protocol is different from other valuable and reliable published protocols for primary mouse RPE isolation13,19,20,24 in that it is simplified with a few steps that are easy to follow. RPE cells are able to be identified by their morphology, pigmentation, and specific RPE markers such as RPE654,5,21. Validation of the purity and contamination of Müller cells was achieved by staining isolated cells with a specific cell marker for Müller cells (vimentin)22. Many techniques have been used to evaluate the integrity of the blood-retinal barrier (BRB) in vitro, such as electron microscope (EM) examination, assessment of tight junction proteins (TJPs), FITIC dextran leakage assay, and transepithelial electrical resistance (TER) measurement that evaluates the physiological function of RPE as a monolayer8,23. RPE cells play an important role in maintaining the outer BRB, and to validate this function, we evaluated tight junction proteins (TJPs) such as Zona ocludin-1 (ZO-1) and cytoskeletal proteins like F-actin as previously published8. TJPs evaluation is a more convenient method for the evaluation of BRB integrity and barrier function which doesn't require expensive equipment that may not be available in all research labs, as opposed to EM evaluation or TER.
In the References section, a 24th reference was added:
- Promsote, W., Makala, L., Li, B., Smith, SB., Singh, N., Ganapathy, V., Pace, B.S., Martin, P. Monomethylfumarate induces γ-globin expression and fetal hemoglobin production in cultured human retinal pigment epithelial (RPE) and erythroid cells, and in intact retina. Invest Ophthalmol Vis Sci. 55(8):5382-93 (2014).
Protocol
An erratum was issued for: Isolation of Primary Mouse Retinal Pigmented Epithelium Cells. The Discussion and References sections were updated.
The third paragraph of the Discussion section was updated from:
This protocol is different from other valuable and reliable published protocols for primary mouse RPE isolation13,19,20 in that it is simplified with a few steps that are easy to follow. RPE cells are able to be identified by their morphology, pigmentation, and specific RPE markers such as RPE654,5,21. Validation of the purity and contamination of Müller cells was achieved by staining isolated cells with a specific cell marker for Müller cells (vimentin)22. Many techniques have been used to evaluate the integrity of the blood-retinal barrier (BRB) in vitro, such as electron microscope (EM) examination, assessment of tight junction proteins (TJPs), FITIC dextran leakage assay, and transepithelial electrical resistance (TER) measurement that evaluates the physiological function of RPE as a monolayer8,23. RPE cells play an important role in maintaining the outer BRB, and to validate this function, we evaluated tight junction proteins (TJPs) such as Zona ocludin-1 (ZO-1) and cytoskeletal proteins like F-actin as previously published8. TJPs evaluation is a more convenient method for the evaluation of BRB integrity and barrier function which doesn't require expensive equipment that may not be available in all research labs, as opposed to EM evaluation or TER.
to:
This protocol is different from other valuable and reliable published protocols for primary mouse RPE isolation13,19,20,24 in that it is simplified with a few steps that are easy to follow. RPE cells are able to be identified by their morphology, pigmentation, and specific RPE markers such as RPE654,5,21. Validation of the purity and contamination of Müller cells was achieved by staining isolated cells with a specific cell marker for Müller cells (vimentin)22. Many techniques have been used to evaluate the integrity of the blood-retinal barrier (BRB) in vitro, such as electron microscope (EM) examination, assessment of tight junction proteins (TJPs), FITIC dextran leakage assay, and transepithelial electrical resistance (TER) measurement that evaluates the physiological function of RPE as a monolayer8,23. RPE cells play an important role in maintaining the outer BRB, and to validate this function, we evaluated tight junction proteins (TJPs) such as Zona ocludin-1 (ZO-1) and cytoskeletal proteins like F-actin as previously published8. TJPs evaluation is a more convenient method for the evaluation of BRB integrity and barrier function which doesn't require expensive equipment that may not be available in all research labs, as opposed to EM evaluation or TER.
In the References section, a 24th reference was added:
- Promsote, W., Makala, L., Li, B., Smith, SB., Singh, N., Ganapathy, V., Pace, B.S., Martin, P. Monomethylfumarate induces γ-globin expression and fetal hemoglobin production in cultured human retinal pigment epithelial (RPE) and erythroid cells, and in intact retina. Invest Ophthalmol Vis Sci. 55(8):5382-93 (2014).
Disclosures
No conflicts of interest declared.
