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Testing method for eliminating fluorescence internal filtration effects by using corrector formula

A test method and fluorescence technology, which is applied in the directions of fluorescence/phosphorescence, color/spectral characteristic measurement, material excitation analysis, etc., can solve cumbersome problems, achieve the effects of simple operation, time saving, and great promotion and application value

Inactive Publication Date: 2012-06-13
SHANDONG UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

This method needs to prepare the solution of the system to be tested and the components in the system to be tested separately, which is cumbersome.

Method used

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  • Testing method for eliminating fluorescence internal filtration effects by using corrector formula
  • Testing method for eliminating fluorescence internal filtration effects by using corrector formula
  • Testing method for eliminating fluorescence internal filtration effects by using corrector formula

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Put the reference absorption cell and the fluorescence cell used to measure the absorption value (spectrum) of the fluorescent sample into ultrapure water, mark one side of the fluorescence cell, make the marked side face the light source side, and set the UV-Vis The scanning wavelength range of the spectrophotometer, so that it includes the range of the excitation wavelength and the emission wavelength (pre-examined) during the fluorescence test of the sample to be tested, for baseline correction; prepare bovine serum albumin (BSA) solution, tetracycline (TC) solution and a mixture of the two (C BSA =1×10 -6 mol / L, C TC =5×10 -5 mol / L, with a phosphate buffer solution to control the pH to be 7.4), put the prepared sample to be tested in the fluorescence cell, with the marked side facing the light source side, and measure the fluorescence emission spectrum of the BSA+TC mixed solution (excitation Wavelength = 278nm, emission wavelength range 280-500nm, take a data po...

Embodiment 2

[0023] Similar to the steps in Example 1, baseline correction is performed first. Prepare catalase (CAT) solution, oxytetracycline (OTC) solution and their mixed system (C CAT =1.5×10 -6 mol / L, C OTC =5×10 -5 mol / L, with phosphate buffer to control the pH to 7.4). Fluorescence spectrum (excitation wavelength=274nm, emission wavelength=283-490nm, temperature is 293K) and absorption spectrum (absorption spectrum measurement range is 273-490nm) and fluorescence spectrum of CAT+OTC mixed solution measured by the technical scheme described in embodiment 1 and Emission spectra were corrected. The fluorescence spectrum before correction is Figure 4 In the curve c, the fluorescence spectrum corrected by the technical scheme of the present invention is Figure 4 Curve a in.

[0024] Contrast experiment: the fluorescent spectrum after adopting the routine correction procedure identical with example 1 correction is curve b( Figure 4 ).

[0025] The peak deviation between curve...

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Abstract

The invention discloses a testing method for eliminating fluorescence internal filtration effects by using a corrector formula, which comprises the following steps of: firstly, placing an absorption cell as a reference and a fluorescent cell for testing an absorption value of a fluorescence sample into an ultra-pure water scanning base line; then carrying out fluorescence testing on a system sample to be tested to obtain a fluorescence value Fobs; placing the fluorescent cell which holds a sample to be tested into an ultraviolet visible spectrophotometer for scanning and summing the absorption values corresponding to the parts of excitation wavelength and emission wavelength to obtain an absorption value A; and substituting into the formula Fcor=Fobs10A / 2 to obtain Fcor which is the corrected fluorescence value. In need of preparing a system solution to be tested and no need of preparing and detecting each component solution, the invention has the characteristics of time saving, drug saving, convenience for operation, economy and practicability, correctness and feasibility, and particularly, greatly improves the work efficiency under the condition of more samples and has larger popularization application values.

Description

technical field [0001] The invention relates to a test method for eliminating the fluorescence inner filter effect. Background technique [0002] After some substances are excited by light of a certain wavelength, within a very short time (10 -8 seconds) emits light with a wavelength greater than the excitation wavelength, which is called fluorescence. Fluorescence technology is widely used in biochemistry and molecular biology research, mainly including the following aspects: qualitative and quantitative determination of substances, research on the physical and chemical properties of biological macromolecules and their molecular structures and conformations, using fluorescence lifetime, Quantum yield and other parameters to study energy transfer phenomena in biological macromolecules. [0003] In the specific application process, if the measured system has absorption at the fluorescence excitation and emission wavelengths, it will weaken the fluorescence intensity of the ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/64G01N21/31G01N21/33
Inventor 刘汝涛池振兴赵星辰
Owner SHANDONG UNIV
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