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Single cell RT-PCR analysis of tyrosine kinase receptor expression in adult rat retinal ganglion cells isolated by retinal sandwiching

Lindqvist, Niclas LU ; Vidal-Sanz, Manuel and Hallböök, Finn (2002) In Brain Research Protocols 10(2). p.75-83
Abstract

We describe a protocol for analysis of gene expression in single, acutely dissociated adult rat retinal ganglion cells using RT-PCR. Retrograde tracing of retinal ganglion cells from the superior colliculi was conducted using Fluorogold. Retinas were dissected and ganglion cells isolated using retinal layer separation (sandwiching). Single, fluorescently labelled retinal ganglion cells were aspirated using a micropipette and used for PCR. Two PCR protocols are described where single cell cDNA was analysed for TrkB and GAPDH or TrkB, TrkC, Ret, Met, ErbB2 and Beta-actin by multiplex-PCR. All five tyrosine kinase receptors were amplified from single retinal ganglion cells. The method will prove useful for the molecular characterization of... (More)

We describe a protocol for analysis of gene expression in single, acutely dissociated adult rat retinal ganglion cells using RT-PCR. Retrograde tracing of retinal ganglion cells from the superior colliculi was conducted using Fluorogold. Retinas were dissected and ganglion cells isolated using retinal layer separation (sandwiching). Single, fluorescently labelled retinal ganglion cells were aspirated using a micropipette and used for PCR. Two PCR protocols are described where single cell cDNA was analysed for TrkB and GAPDH or TrkB, TrkC, Ret, Met, ErbB2 and Beta-actin by multiplex-PCR. All five tyrosine kinase receptors were amplified from single retinal ganglion cells. The method will prove useful for the molecular characterization of adult retinal ganglion cells.

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author
publishing date
type
Contribution to journal
publication status
published
keywords
Fluorogold (FG), Polymerase chain reaction (PCR), Retinal ganglion cell (rgc), Reverse transcription (RT)
in
Brain Research Protocols
volume
10
issue
2
pages
9 pages
publisher
Elsevier
external identifiers
  • scopus:0036808209
ISSN
1385-299X
DOI
10.1016/S1385-299X(02)00184-8
language
English
LU publication?
no
id
9fae4c88-f5fd-4714-b60e-01822ac6c734
date added to LUP
2017-06-02 15:18:55
date last changed
2017-06-07 07:56:53
@article{9fae4c88-f5fd-4714-b60e-01822ac6c734,
  abstract     = {<p>We describe a protocol for analysis of gene expression in single, acutely dissociated adult rat retinal ganglion cells using RT-PCR. Retrograde tracing of retinal ganglion cells from the superior colliculi was conducted using Fluorogold. Retinas were dissected and ganglion cells isolated using retinal layer separation (sandwiching). Single, fluorescently labelled retinal ganglion cells were aspirated using a micropipette and used for PCR. Two PCR protocols are described where single cell cDNA was analysed for TrkB and GAPDH or TrkB, TrkC, Ret, Met, ErbB2 and Beta-actin by multiplex-PCR. All five tyrosine kinase receptors were amplified from single retinal ganglion cells. The method will prove useful for the molecular characterization of adult retinal ganglion cells.</p>},
  author       = {Lindqvist, Niclas and Vidal-Sanz, Manuel and Hallböök, Finn},
  issn         = {1385-299X},
  keyword      = {Fluorogold (FG),Polymerase chain reaction (PCR),Retinal ganglion cell (rgc),Reverse transcription (RT)},
  language     = {eng},
  number       = {2},
  pages        = {75--83},
  publisher    = {Elsevier},
  series       = {Brain Research Protocols},
  title        = {Single cell RT-PCR analysis of tyrosine kinase receptor expression in adult rat retinal ganglion cells isolated by retinal sandwiching},
  url          = {http://dx.doi.org/10.1016/S1385-299X(02)00184-8},
  volume       = {10},
  year         = {2002},
}