Fluorometer - Revision history

Cmditradmin: /* Significance */

2011-10-12T15:35:28Z

Significance

← Older revision		Revision as of 08:35, 12 October 2011
Line 78:		Line 78:
	φ<sub>reference</sub> = 0.87)		φ<sub>reference</sub> = 0.87)

	:<math>\~~phi~~ = \phi_{reference} \frac {F_{sample} / A_{sample}} {F_{reference}/ A_{reference}} \left( \frac {n_{sample}} {n_{reference}} \right) ^2\,\!</math>		:<math>\phi_{sample} = \phi_{reference} \frac {F_{sample} / A_{sample}} {F_{reference}/ A_{reference}} \left( \frac {n_{sample}} {n_{reference}} \right) ^2\,\!</math>


	:<math>\~~phi~~ = 0.87 * \frac {1.289} {1.316} * \left( \frac {1.3288} {1.4266} \right) ^2\,\!</math>		:<math>\phi_{sample} = 0.87 * \frac {1.289} {1.316} * \left( \frac {1.3288} {1.4266} \right) ^2 = 8.87 * 0.979* 0.868= 0.74\,\!</math>

	=== Operation ===		=== Operation ===

Cmditradmin at 16:33, 5 October 2011

2011-10-05T16:33:33Z

← Older revision		Revision as of 09:33, 5 October 2011
Line 1:		Line 1:
			<table id="toc" style="width: 100%">
			<tr>
			<td style="text-align: center; width: 33%">[[Main_Page#Research Equipment, Devices and Techniques\|Return to Research Tool Menu]]</td>

			</tr>
			</table>

	=== Background ===		=== Background ===
	[[Image:Fluorometer_layout.png\|thumb\|400px\|Internal layout of fluorometer components- Fluorolog-3, model FL3-12 (Horiba Jobin Yvon) ]]		[[Image:Fluorometer_layout.png\|thumb\|400px\|Internal layout of fluorometer components- Fluorolog-3, model FL3-12 (Horiba Jobin Yvon) ]]

Smhunter at 20:08, 28 July 2011

2011-07-28T20:08:08Z

← Older revision		Revision as of 13:08, 28 July 2011
Line 78:		Line 78:
	=== Operation ===		=== Operation ===

	{{#ev:youtube\|~~qNegKeDrjE4~~}}		{{#ev:youtube\|5edJlvxcxrY}}

	=== External Links ===		=== External Links ===
	[[Wikipedia:Fluorescence spectroscopy]]		[[Wikipedia:Fluorescence spectroscopy]]

Cmditradmin: /* Operation */

2011-06-22T21:03:55Z

Operation

← Older revision		Revision as of 14:03, 22 June 2011
Line 78:		Line 78:
	=== Operation ===		=== Operation ===

			{{#ev:youtube\|qNegKeDrjE4}}

	=== External Links ===		=== External Links ===
	[[Wikipedia:Fluorescence spectroscopy]]		[[Wikipedia:Fluorescence spectroscopy]]

Cmditradmin: /* Background */

2011-06-16T16:32:04Z

Background

← Older revision		Revision as of 09:32, 16 June 2011
Line 1:		Line 1:
	=== Background ===		=== Background ===
	[[Image:Fluorometer_layout.png\|thumb\|400px\|Internal layout of fluorometer components ]]		[[Image:Fluorometer_layout.png\|thumb\|400px\|Internal layout of fluorometer components- Fluorolog-3, model FL3-12 (Horiba Jobin Yvon) ]]
	A fluorometer consists of an excitation monochromator which directs light of specific wavelength on a sample. A fluorescent sample produces and emission spectrum that is measured through a monochromator connected to a photomultiplier detector similar to spectrophotometer.		A fluorometer consists of an excitation monochromator which directs light of specific wavelength on a sample. A fluorescent sample produces and emission spectrum that is measured through a monochromator connected to a photomultiplier detector similar to spectrophotometer.

Cmditradmin: /* Background */

2011-02-28T22:41:38Z

Background

← Older revision		Revision as of 15:41, 28 February 2011
Line 1:		Line 1:
	=== Background ===		=== Background ===
	[[Image:Fluorometer_layout.png\|thumb\|400px\|Internal layout of fluorometer components ]]		[[Image:Fluorometer_layout.png\|thumb\|400px\|Internal layout of fluorometer components ]]
	A fluorometer consists of an excitation monochromator which directs light of specific wavelength on a sample. A fluorescent sample produces and emission spectrum that is measured through a monochromator connected to a photomultiplier detector similar to spectrophotometer~~. A typical fluorometer is able to create both an absorption an emission spectrum for a liquid sample~~.		A fluorometer consists of an excitation monochromator which directs light of specific wavelength on a sample. A fluorescent sample produces and emission spectrum that is measured through a monochromator connected to a photomultiplier detector similar to spectrophotometer.

	=== Significance ===		=== Significance ===

Cmditradmin: /* Background */

2011-02-28T22:25:15Z

Background

← Older revision		Revision as of 15:25, 28 February 2011
Line 1:		Line 1:
	=== Background ===		=== Background ===
	[[Image:Fluorometer_layout.png\|thumb\|~~300px~~\|Internal layout of fluorometer components ]]		[[Image:Fluorometer_layout.png\|thumb\|400px\|Internal layout of fluorometer components ]]
	A fluorometer consists of an excitation monochromator which directs light of specific wavelength on a sample. A fluorescent sample produces and emission spectrum that is measured through a monochromator connected to a detector similar to spectrophotometer. A typical fluorometer is able to create both an absorption an emission spectrum for a liquid sample.		A fluorometer consists of an excitation monochromator which directs light of specific wavelength on a sample. A fluorescent sample produces and emission spectrum that is measured through a monochromator connected to a photomultiplier detector similar to spectrophotometer. A typical fluorometer is able to create both an absorption an emission spectrum for a liquid sample.

	=== Significance ===		=== Significance ===

Cmditradmin: /* Background */

2011-02-28T22:18:04Z

Background

← Older revision		Revision as of 15:18, 28 February 2011
Line 1:		Line 1:
	=== Background ===		=== Background ===
			[[Image:Fluorometer_layout.png\|thumb\|300px\|Internal layout of fluorometer components ]]
			A fluorometer consists of an excitation monochromator which directs light of specific wavelength on a sample. A fluorescent sample produces and emission spectrum that is measured through a monochromator connected to a detector similar to spectrophotometer. A typical fluorometer is able to create both an absorption an emission spectrum for a liquid sample.

	=== Significance ===		=== Significance ===

Cmditradmin: /* Significance */

2011-02-28T22:07:01Z

Significance

← Older revision		Revision as of 15:07, 28 February 2011
Line 6:		Line 6:
	'''Fluorescence quantum yield determination using relavitive method'''		'''Fluorescence quantum yield determination using relavitive method'''

	One significant use of the fluorometer (or fluorimeter) is the determination of the fluorescence quantum yield. This is done using a relative method based on a reference compound of known quantum yield. The unknown sample and the reference sample are measured at the same excitation wavelengths and measurement conditions. The wavelength-integrated ~~flourescent~~ intensity of both materials are then used in the calculation:		One significant use of the fluorometer (or fluorimeter) is the determination of the fluorescence quantum yield. This is done using a relative method based on a reference compound of known quantum yield. The unknown sample and the reference sample are measured at the same excitation wavelengths and measurement conditions. The wavelength-integrated fluorescent intensity of both materials are then used in the calculation:

	:<math>\phi = \phi_{reference} \frac {F_{sample} / A_{sample}} {F_{reference}/ A_{reference}} \left( \frac {n_{sample}} {n_{reference}} \right) ^2\,\!</math>		:<math>\phi = \phi_{reference} \frac {F_{sample} / A_{sample}} {F_{reference}/ A_{reference}} \left( \frac {n_{sample}} {n_{reference}} \right) ^2\,\!</math>
Line 24:		Line 24:
	The intensity of the excitation beam should be almost constant along excitation beam. The fluorescence signal is proportional to intensity of excitation beam and the intensity has to be the same for sample and reference.		The intensity of the excitation beam should be almost constant along excitation beam. The fluorescence signal is proportional to intensity of excitation beam and the intensity has to be the same for sample and reference.

	Typically: A ≤ 0.02 over 1 cm ~~pathlength~~		Typically: A ≤ 0.02 over 1 cm path length

	From entrance face to center of cuvette: A = 0.01		From entrance face to center of cuvette: A = 0.01
Line 32:		Line 32:
	Therefore the intensity has changed only by 2%		Therefore the intensity has changed only by 2%

	~~Depending~~ on absorption spectrometer used, measurement of A in this range may not be accurate enough. How to proceed?		However, depending on absorption spectrometer used, measurement of A in this range may not be accurate enough. How to proceed?
	*Measure A on higher concentration solution		*Measure A on higher concentration solution
	*Dilute solution by (accurately) known factor		*Dilute solution by (accurately) known factor
Line 44:		Line 44:

	In this test, we are using: λ<sub>exc</sub> = 350 nm		In this test, we are using: λ<sub>exc</sub> = 350 nm


	'''Repeated measurements'''		'''Repeated measurements'''

	Its always a good idea to have multiple data points! Prepare multiple dilutions and then measure fluorescence emission spectrum of each solution. Determine the slope of the line F/A for sample and reference. Deviations from linearity could indicate that emission was affected by reabsorption		Its always a good idea to have multiple data points! Prepare multiple dilutions and then measure fluorescence emission spectrum of each solution. Determine the slope of the line F/A for sample and reference. Deviations from linearity could indicate that emission was affected by reabsorption

			[[Image:Fluor_dilution.png\|thumb\|300px\| Fluorescence intensity for various dilutions]]

	'''Reabsorption'''		'''Reabsorption'''
Line 57:		Line 58:
	[[Image:Solvent_subtraction.png\|thumb\|500px\| ]]		[[Image:Solvent_subtraction.png\|thumb\|500px\| ]]
	Detectors and gratings do not have the same efficiency at all wavelengths. Therefore the results need to be corrected by a factor that accounts for wavelength response of the instrument. The contribution of the solvent (Raman scattering) and noise (dark counts) should also be subtracted.		Detectors and gratings do not have the same efficiency at all wavelengths. Therefore the results need to be corrected by a factor that accounts for wavelength response of the instrument. The contribution of the solvent (Raman scattering) and noise (dark counts) should also be subtracted.
			<br clear='all'>

	'''Sample Calculation'''		'''Sample Calculation'''

Cmditradmin: /* Significance */

2011-02-28T21:40:46Z

Significance

← Older revision		Revision as of 14:40, 28 February 2011
Line 59:		Line 59:

	'''Sample Calculation'''		'''Sample Calculation'''

	F/A values n		F/A values n