Endy:Victor3 plate reader/detection limits and linear range

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Current revision (18:13, 6 January 2006) (view source)
 
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==Absorbance measurements==
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[[Image:TableSpec to Plate Reader.png|thumb|200px|right|Instrument reading for a serial dilution of cell culture]]
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For the OD600 label, the machine appears to respond in a linear fashion over the range of absorbances shown in this figure.  More details of the experiment can be found here.  Better data over a larger range will follow shortly.
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==Fluorescence measurements==
Recent experiments have raised the question of what range of concentrations can we expect the plate reader to give linear results.  Chris Quinlan ([http://www.perkinelmer.com of Perkin Elmer]) provided the following information about the plate reader's detection limits.
Recent experiments have raised the question of what range of concentrations can we expect the plate reader to give linear results.  Chris Quinlan ([http://www.perkinelmer.com of Perkin Elmer]) provided the following information about the plate reader's detection limits.
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*Fluorescein MDL (Minimum detectable level) - 2fMoles/well.  Using 200<math>\mu</math>l per well this corresponds to 10pM
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*Fluorescein MDL (Minimum detectable level) - 2fMoles/well.  Using 200<math>\mu</math>l per well this corresponds to 10pM.
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*([http://www.perkinelmer.com Perkin Elmer]) believe the machine has a five decade linear range.  This means concentrations up to <math>\mu</math>M should be read linearly.
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*([http://www.perkinelmer.com Perkin Elmer]) believe the machine has a five decade linear range.  This means concentrations up to <math>\mu</math>Molar should be read linearly.
*For higher concentrations, use a smaller volume.
*For higher concentrations, use a smaller volume.
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*The emission of Fluorescein relative to GFP is not yet known.  We also do not know how the emission of purified fluorophores relates to the emission of fluorophores ''in vivo''
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*The emission of Fluorescein relative to GFP is not yet known.  We also do not know how the emission of purified fluorophores relates to the emission of fluorophores ''in vivo''.
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*More to follow on this hopefully.

Current revision

Absorbance measurements

Instrument reading for a serial dilution of cell culture
Instrument reading for a serial dilution of cell culture

For the OD600 label, the machine appears to respond in a linear fashion over the range of absorbances shown in this figure. More details of the experiment can be found here. Better data over a larger range will follow shortly.


Fluorescence measurements

Recent experiments have raised the question of what range of concentrations can we expect the plate reader to give linear results. Chris Quinlan (of Perkin Elmer) provided the following information about the plate reader's detection limits.

  • Fluorescein MDL (Minimum detectable level) - 2fMoles/well. Using 200μl per well this corresponds to 10pM.
  • (Perkin Elmer) believe the machine has a five decade linear range. This means concentrations up to μMolar should be read linearly.
  • For higher concentrations, use a smaller volume.
  • The emission of Fluorescein relative to GFP is not yet known. We also do not know how the emission of purified fluorophores relates to the emission of fluorophores in vivo.
  • More to follow on this hopefully.
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