Quantitative dual-dye photometric method for determining dilution impact

• US 7,998,747 B2
• Filed: 09/13/2007
• Issued: 08/16/2011
• Est. Priority Date: 09/15/2006
• Status: Active Grant

• Current Assignee: Artel Incorporated (Conagra Brands, Inc.)
• Original Assignee: Artel Incorporated (Conagra Brands, Inc.)
• Inventors: Curtis, Richard H., Bradshaw, John Thomas, McNally, Ceara, Rogers, Alex L., Albert, Keith J., Knaide, Tanya R.
• Primary Examiner(s): Soderquist; Arlen
• Assistant Examiner(s): XU, XIAOYUN
• Application Number:
  US11/854,594
• Publication Number:
  US 20080070317A1
• Time in Patent Office: 1,433 Days
• Field of Search: None
• US Class Current: 436/164
• CPC Class Codes: G01N 21/31 Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry G01N21/72 takes precedence
• CPC Class Codes: Y10T 436/25625 Dilution
• CPC Class Codes: Y10T 436/2575 Volumetric liquid transfer

Abstract

The invention provides ways to determine the impact of diluting a solution wherein the diluting may be carried out for any of a variety of purposes. In one embodiment, the method enables accurate volume dispensation calculations independent of meniscus shape. In another embodiment, the method enables accurate determination of plate washing efficiency. In yet another embodiment, the method enables the accurate determination of dilution ratio over a plurality of dilution steps. The methods described may be carried out using one or more systems arranged to perform the steps. A kit of the invention includes instructions for carrying out the steps of the methods and, optionally, one or more solutions suitable for conducting photometric measurements.

Claims

1. A method of determining the dilution achieved in carrying out a dilution protocol using a sample solution and a diluent, wherein the sample solution includes a first chromophore that absorbs light at a first wavelength and a known concentration of a second chromophore that absorbs light at a second wavelength, the method comprising the steps of:

a. adding to a first vessel of a first set of a plurality of vessels a target volume of a first sample solution, wherein the first sample solution includes a first known concentration of the first chromophore and a known concentration of the second chromophore, wherein each of the first chromophore and the second chromophore has an absorbance maximum, the absorbance maximum of the first chromophore minimally overlapping with the absorbance maximum of the second chromophore;

b. adding to a first vessel of a second set of the plurality of vessels a target volume of a second sample solution including a second known concentration of the first chromophore and the known concentration of the second chromophore, wherein the second known concentration of the first chromophore of the second sample solution is higher than the first known concentration of the first chromophore of the first sample solution, and wherein the target volume of the second sample solution added to the first vessel of the second set is substantially equivalent to the target volume of the first sample solution added to the first vessel of the first set;

c. carrying out a first dilution protocol step comprising;

i. mixing into the first vessel of the first set a first target volume of a diluent, wherein the diluent includes the second chromophore at a concentration substantially equivalent to the known concentration of the second chromophore in the first sample solution and the second sample solution;

ii. mixing into the first vessel of the second set the first target volume of the diluent; and

iii. measuring the absorbance of the first chromophore at the first wavelength and the absorbance of the second chromophore at the second wavelength in the first vessel of the first set;

d. carrying out a second dilution protocol step comprising;

i. transferring a target volume of the mixture of the first sample solution and the diluent of the first vessel of the first set to a second vessel of the first set;

ii. mixing a second target volume of the diluent into the second vessel of the first set;

iii. transferring a target volume of the mixture of the second sample solution and the diluent of the first vessel of the second set to a second vessel of the second set;

iv. mixing the second target volume of the diluent into the second vessel of the second set; and

v. measuring the absorbance of the first chromophore at the first wavelength and the absorbance of the second chromophore at the second wavelength in the second vessel of the second set; and

e. calculating a dilution ratio of the dilution protocol based on the absorbance measurements made in steps c.iii and d.v, wherein the dilution ratio represents the extent of dilution occurring between the first dilution protocol step and the second dilution protocol step.

2. The method of claim 1 wherein the calculation of the dilution ratio involves using the equation:

3. The method of claim 1 further comprising the steps of:

a. repeating X more times the dilution protocol steps of i) transferring the sample solution and the diluent mixture to subsequent vessels for each of the first and second sets;

ii) mixing a target volume of the diluent into the subsequent vessels and iii) measuring the absorbances of the first and second chromophores, wherein X is ≧

1, such that the last dilution protocol step is dilution protocol step n and a preceding dilution protocol step is dilution protocol step m, and wherein vessels of the first set and the second set containing the mixture of the sample solution and the diluent after completing dilution protocol step m are represented as vessel m and vessels of the first and second set containing the mixture of the sample solution and the diluent after completing dilution protocol step n are represented as vessel n; and

b. calculating a dilution ratio of the dilution protocol based on the absorbance measurements made after completing dilution protocol step m and dilution protocol step n, wherein the dilution ratio represents the extent of dilution occurring between dilution protocol step m and dilution protocol step n.

4. The method of claim 3 wherein the calculation of the dilution ratio involves using the equation:

5. The method of claim 1 wherein the first target volume of the diluent and the second target volume of the diluent are not equal.