MXPA97007500A - Method for the calibration of an oximeter and to stop result - Google Patents

Abstract

The present invention relates to a method for determining the performance of an absorbency spectrometer, the method comprising: a. measure the absorbance spectrum of a quality control sample b. fix the sample spectrum measured with the normal spectrum of the quality control sample to obtain a spectrum of better fixation and an apparent concentration of the quality control sample; evaluate the quality of the fixation

Description

METHOD FOR THE CALIBRATION OF AN OXIMETER AND TO REPORT RESULTS FIELD OF THE INVENTION This invention relates to the field of evaluation of the operation of spectrophotometers.

BRIEF DESCRIPTION OF THE RELATED ART Infrared visible ultraviolet absorbance spectroscopy (UV-VIS-IR) has become an invaluable tool in medical diagnosis and analytical chemistry. For example, CO-oximeters measure the concentrations of various components of hemoglobin or fractions in physiological blood using absorption spectroscopy. The CO-oximeters perform this function by measuring the VIS-IR absorbance spectrum of a blood sample and determining the best fixation of the known blood component spectra. Therefore, it is easily appreciated that it is important to be able to quickly and accurately determine the performance of instruments. Existing methods measure the absorbance spectrum of quality control products. The QC products specific for blood analysis are typically red dye-based samples constructed to simulate the blood spectrum. In addition to a red dye, they sometimes contain certain amounts of oxygen, carbon dioxide, and electrolytes, at a set pH, to determine the performance of gas and electrolyte instruments for blood. It is very difficult to build QC products that have an absorbance spectrum that closely resembles that of physiological blood. Normal blood component spectra are used to obtain the best fixation to the spectrum of the measured QC product. By varying the contributions of each blood component spectrum, a better fixation to the QC spectrum is obtained. But since even the ideal QC spectrum does not resemble that of physiological blood, the resulting lower fixation deviates from the observed QC spectrum, even when the instrument is working perfectly. In addition, errors induced by the instrument are not reported in terms easily understood by the ordinary technician who operates the instrument. For example, current methods report the fraction of major blood components (as determined from the relative contribution of each spectrum of the blood component to the spectrum of best fixation). Usually a fractional concentration of a blood component is negative. Consequently, improved methods are desired to determine and report the operation of the instrument.

COMPENDIUM OF THE INVENTION The present invention provides methods for determining and reporting the performance of VIS-I R spectrophotometers as used to measure the concentration of hemoglobin components or fractions in blood samples. In particular, the method is used to determine the performance of CO-oximeters. In one aspect of the invention, the methods comprise measuring the absorbance spectrum of a quality control product sample (QC product) and comparing it with the known normal spectrum for the same QC product. The normal known spectrum of the QC product is obtained from a well-calibrated, highly accurate spectrophotometer under controlled conditions and electronically stored in the instrument by the manufacturer for future comparison. The QC sample spectrum measured in commercial analytical instruments will differ from the normal spectrum mainly due to several instrumental errors. The normal spectrum is mathematically fixed to the measured spectrum, producing an apparent concentration of the QC. The difference between the measured spectrum and the normal spectrum of best fixation (the "error spectrum") and the variation of the QC concentration from the normal concentration QC, is a measure of the instrumental error. An important use of the above method is to calibrate the operation of spectrophotometers used for analytical determinations, including the concentration of various blood components. This method to determine the instrumental performance is easily conducted by the technician in the laboratory, and involves simply measuring the absorbance spectrum of a QC sample. The instrument can be programmed to automatically carry out the previous procedure and report the instrumental error. If the observed instrumental error is too great, the instrument may require service before analyzing test samples. This method is distinguished from the prior art methods since the closely measured spectrum resembles the normal spectrum to which it is compared. In addition, this method also allows the results to be reported in terms easily recognized and easily appreciated by the technician. In another aspect of the present invention, the concentration of the QC obtained from the mathematical fixation is converted to an apparent total concentration of hemoglobin (Tapp). This is achieved by multiplying the apparent concentration of the QC by a nominal value of the total concentration of hemoglobin (Tnom), which, in one aspect of the invention, is equal to the normal level of hemoglobin found in human blood. Tapp is the total concentration of hemoglobin that could have been determined from the absorption spectrum of a test sample having a hemoglobin concentration of Tnom. This number is easily appreciated by the technician, and, if it differs too much from the nominal concentration of human hemoglobin, it will indicate to the technician that the instrument may require service. In another aspect, the QC spectrum and spectral measurement errors are conveniently translated into blood component concentrations, i.e., hemoglobin and lipid fractions. This allows easy recognition and appreciation of QC results by technicians. This can be done in one of these two ways. The simplest way is to multiply the apparent total concentrations of hemoglobin (Tapp) by the nominal fraction of each component in the blood. The nominal fractions are preferably the fraction of the total concentration of hemoglobin of the particular component (eg, oxygenated hemoglobin) normally found in the physiological blood. This method presents the same information contained in Tapp, but in a different format. The second method, very preferred, to determine and report fractional concentrations of blood component, uses a mathematical analysis to determine the errors in concentrations of the main blood components due to instrumental errors. This method determines the values of blood component concentrations that could give rise to the error spectrum. Several blood component spectra are mathematically combined to give the best fixation to the error spectrum. The contributions of each blood component spectrum directly produce the concentration of the blood component. These concentrations are the deviations (due to instrumental error) of the actual concentrations of these components that one might observe if a real blood sample were analyzed. The addition of these concentrations to the nominal concentrations of the blood components, produces the concentrations of the blood components that could have been observed if a blood sample, having the nominal concentrations of the blood components, was analyzed in the instrument. In a preferred embodiment, the nominal values of the blood component concentrations are set at those values observed in normal human blood. The importance of the results can then be easily appreciated by the person skilled in the art. If the errors go beyond acceptable limits, the service may be required. The method herein provides an improved ability to determine instrument performance, since the measured QC sample spectrum closely resembles the known, normal spectrum. The result is that the errors measured relate more directly to the instrument errors. Concomitantly, the difficult task of formulating stable QC products that resemble blood spectra is eliminated. In addition, the same QC product can be used in instruments using different wavelengths for the measurement of parameters and retain the ability to report the results at the same nominal value. These nominal values can advantageously be set to any desired value.

The method herein also provides the ability to classify the concentration of blood components to better reflect anticipated instrument variations when performing physiological blood samples. The foregoing merely summarizes certain aspects of the invention and is not intended, nor should it be constructed, to limit the invention in any way.

BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 presents three QC spectra of the three batches of the QC product of level 1 of Example 1, CERTAIN® ELITE. Figure 2 presents the spectra of the main absorption blood components. Figure 3 presents the spectra that are the difference between the QC spectra of Figure 1 and the best least squares fix for those spectra that use the blood component spectra of Figure 2. Figure 4 presents the error spectra, is say, the difference between the measured QC spectra of Figure 1 and the best least squares fixation of normal QC, which in this case is the average of the three QC spectra of Figure 1. Figure 5 presents the spectra that are the difference between the error spectrum of Figure 4 and the best least-squares binding to that the spectra used by the blood component spectra of Figure 2.

DETAILED DESCRIPTION OF THE PREFERRED MODALITIES The present invention provides a new method for determining the level of performance of an absorbance spectrophotometer. As used herein, the terms "absorbance spectrometer", "spectrometer", and their equivalents, mean any machine or instrument capable of measuring the absorbance spectrum of a liquid sample. The methods are particularly useful for determining the performance of CO-oximeters. The methods comprise measuring the absorbance spectrum of a quality control sample (QC) and evaluating the quality of the binding to the known spectrum, normal for that QC product. The difference between the two is a measure of the operation of the spectrophotometer. This method is distinguished from prior art methods, which compare measured QC spectra with a normal blood spectrum. Since the QC spectra do not closely resemble the blood spectra, the differences observed in the measured QC spectrum and a normal blood spectrum are rather large (Figure 3) and increase due to both variations in instrument performance and by the inherent differences in the two spectra. The methods of the present are improved by this greatly reducing or eliminating the inherent difference between the measured QC sample spectrum and the normal spectrum with which it is compared. Figures 3 and 4. A disadvantage of the prior art is that blood component concentrations obtained from QC spectra usually have little or no relationship to actual values in the blood, usually appearing as negative concentrations or concentrations greater than 100% . The methods of the present are improved with respect to the prior art, in that the instrumental error can be reported in terms of blood component concentrations that are comparable with actual values in the blood, and thus easily understood by the skilled artisan. technique. The method of the invention is based on the well-known law of Beer-Lambert: A? = q? • c • x (1) where A is the absorbance at the wavelength?, Q? Is the extinction coefficient at wavelength? for the absorber, c is the concentration of the absorber and x is the path length of the sample, through which the measurement lz beam passes. The path length x is constant within the spectrophotometer (typically expressed in cm) and will henceforth be included in q. If there are "n" components in the sample, Beer-Lambert's law can be expressed in linear algebraic terms such as: A = q • c, (2) where A is a vector whose "m" elements A? ¡Are the absorbances at "m" discrete wavelengths? ¡, Q is a "mxn" matrix whose elements qj are the extinction coefficients of the component "j" at the wavelength? ¡, and c is a vector, each of whose "n" elements Cj is the concentration of the component "j". In the present invention, the absorbance spectrum (Amßaß) of a QC sample is measured (Fig. 1). The QC sample has a known group of extinction coefficients q, as determined by measuring the absorbance spectrum or the normal spectrum (with a cstd concentration) in a calibrated spectrophotometer. During practice, a spectrum of normal bsorbancy (Astd) of a QC product will be recorded in a high-precision spectrophotometer and the concentration (cst <J) will arbitrarily be set to unity, so that q = Astd - Alternatively, the standard QC spectrum can be obtained by averaging two or more QC sample pecters. The QC samples used to determine the operation of the instrument are fabricated to be identical (ie have the same cstd) to the QC product used to obtain A8td. It is observed that the variation in the product specifications of different lots of QC products may impact the analysis, although this variation must be smaller. The vector q is then electronically stored in the instrument by the manufacturer for use in further determinations of the operation of the instrument. The QC spectra will generally differ from the samples (for example, blood samples) that are commonly analyzed in the spectrophotometer. As used herein, the term "analyze" means determining the identity and / or concentration of the component (s) of a sample. To give the most accurate and meaningful indication of the performance of the instrument, the QC sample should preferably be constructed to give a spectrum of absorbance closely resembling samples frequently analyzed in the spectrophotometer. In a preferred embodiment, the QC sample will resemble human blood. The QC products suitable for use in the invention are commercially available, for example, from Ciba Corning Diagnostics Corp. (Medfield, MA) under the trade name CERTAIN® ELITE. The normal QC spectrum, Astd, is fixed to the measured spectrum, Ameas, treating "c" in equation (2) as a variable and determining the value of "c" that results in the best fixation. The result is an apparent concentration of the QC, capp. A variety of potential sources of instrumental error will result in Amßa8 which differs from Astd and, consequently, capp which differs from cstd. Any of the well-known mathematical techniques for setting the spectra can be used to determine the best fixation of A8td to Ameas and from this to capp. See Press et al., Numerical Recipes: The Art of Scientific Computing, Chapter 14, p. 498-546 (Cambridge University Press, Cambridge, 1986). In a preferred embodiment, capp is obtained through the least squares analysis and is given by: capp = (qtq) "1qt.AfTl < (3) See, Noble and Daniel, Applied Linear Algebra, p. 57-65 (Prentice-Hall 1977). The apparent concentration, capp, can then be used to give the absorbance spectrum of best fixation, estimated (in the least squares sense), Aßst: Aßst = q »capp (4) The difference between the absorbance spectra measured and estimated at each wavelength, that is, the" error spectrum "is given by the vector E: E = Aest - Ameas (5) (Figure 4). The vector E is a measure of the deviation of instrumental functioning from the ideal. It is difficult to determine from the same E, however, the level of operation of the instrument and whether the instrument is operating within acceptable levels. In a preferred embodiment of the present invention, E is advantageously related to errors in the measured concentration of blood components (C? Hb) that could result from instrumental errors. C? Hb is a vector that has "k" elements, each being the error in concentration of one of the blood components C? Hb is the vector of blood component concentrations that could give rise to the observed error spectrum, E. C? Hb can be determined in a variety of ways involving adjusting the error spectrum with one or more known blood component spectra by varying the intensities of each blood component spectrum. The relative contributions of each of the blood component spectra are the relative concentrations of each of the components. Preferably, the least squares method is used. If QHb is the "k x m" matrix of the extinction coefficients for the blood components "k" to "m" wavelengths, then C? Hb is given by: C? Hb = (QHbTQHb) '1QHbT »E (6a) C? Hb = W. (HbtQHb) "1QHbT« E (6b) The C? Hb thus obtained will have the same units as QHb, preferably gm / dL The values of C? Hb can be positive (indicating that the instrumental errors could result in measured blood component concentrations that are too high), or negative (indicating that the instrumental error could result in measured blood component concentrations that are too low.) If all the elements of E were with a zero letter, that is, , Amßas = A, td, meaning that the instrument worked ideally, there could be no error in the measurements of the concentrations of the blood components, and all the elements of C? Hb would be zero.The weight or W is a matrix, which classifies the concentration of blood components to better reflect anticipated instrument variations when performing physiological blood samples.The simplest case is when the diagonal elements are n equal to 1 and the others are 0. Equation (6b) provides a transformation of the error concentrations obtained with QC, so that they reflect values that could have been obtained if the sample were oxyhemoglobin. A physiological sample transformation is a matrix that weighs each error fraction to fit a new value. Errors due to instrument inaccuracies, such as spectral displacement, or wavelength, result in characteristic errors for each fraction of hemoglobin, as well as for quality control. The characteristic response of the instrument to such inaccuracies may be predetermined for any specific sample. Since the physiological samples are predominantly oxyhemoglobin, and since the characteristic response of the instrument (due to inaccuracies) will differ for samples other than oxyhemoglobin, it is desirable to compensate for these differences. In another preferred embodiment of the invention, the total concentration of hemoglobin is reported (Tap p). It is the product of the apparent concentration of the measured QC, capp, and the nominal value of the total hemoglobin Tnom: 'a p p -' not m * Ca p p \ ') Tnom can be set at any convenient value. In a preferred embodiment, Tnom = 14 g / dL, a typical value of normal human blood. In this way, if capp = 1 (which is the arbitrary heat chosen for the QC product and could result if Am eas = A8td), the reported total hemoglobin could be equal to that observed in normal human blood. The nominal concentration of the "k" components of blood, "nom -" nom • 'nc (8) where Fnom is the vector of each of its elements Fnom, are the nominal fractions of the blood component "i". preferred embodiment of the present invention the Fnom elements are set for the actual fractions of the blood components observed in human blood.The apparent concentration of blood components that one could observe in the instrument being tested (Cxb.ap), is the Sum of the nominal concentrations (Cnom) and the errors in the blood component concentrations previously calculated, C? Hb: The total calculated concentration of hemoglobin (the scalar, caic) is the sum of the calculated values of the components, that is, the sum of the elements of the vector CHb, aPp- The reported fractions of the Frep blood components are given by: Frßp = Cpb.app/Tcalc (10) The evaluation of one or more of the indications to After the instrumental operation, it will be possible for the technician in the field to determine if it is necessary to service the spectrophotometer or to continue the analysis of the test sample.

That is, one or more of the following points can be observed: 1. the deviation of the quality control spectrum of better fixation (Aßßt) from the measured QC spectrum (A ^ ,,,) or, equivalently, the spectrum of error, E; 2. the deviation of capp from cstd; 4. the deviation of Tapp or Tca? C of Tn0m; 5. the deviation of CHb, aPP from Cn0m; 6. Frep deviation from Fno? «; and 7. any equivalent indication of instrumental performance derived from the comparison of Amßas and Astd. The acceptable limits within which the instrument is required to function will depend on particular circumstances and will vary from application to application. For example, in cases where only rigorous estimates of concentrations are required, a large deviation from any of the above ideal parameters can be tolerated. On the other hand, where extremely precise concentration measurements are required, only small deviations in the above parameters will be acceptable. Typical acceptable values will vary within approximately + 2% of the nominal value. The main components of blood that ordinarily will be used in the methods of the present are reduced hemoglobin (HHb), oxygenated hemoglobin (O2Hb), carboxyhemoglobin (COHb), methemoglobin (MetHb), sulf hemoglobin (SHb), and lipid. The extinction coefficients for blood components based on hemoglobin can be obtained directly or obtained from the literature. See, Zijlstra et al., Clin. Chem. 37 (9), 1633-1638 (1991). The lipid spectrum can be measured through intravenous fat emulsion (eg, the commercially available lipid product, LIPOSYN ™ II) in an aqueous dispersion of about 10% by weight. Any number of wavelengths can be used to measure and set the previous spectra, but at a minimum there must be at least as many wavelengths as variables (ie concentrations) that will be set. The larger the number of sling lengths used, the smaller the contribution that will exist to the random noise, and, therefore, the more accurate the measurements will be. As a practical matter, seven wavelengths can be used to set an error spectrum of 6-component spectra, for example, HHb, O2Hb, COHb, MetHb, SHb, and lipid. Preferably, the spectra are measured and fixed at wavelengths, at which the QC products have at least a moderate level of absorbance. The following examples are offered for illustrative purposes only and are not intended, nor should be constructed, to limit the invention in any way.

EXAMPLES EXAMPLE 1 The methods of the present invention were applied to three different batches of three different QC CERTAIN® ELITE products (Ciba Corning Diagnostics, Medfield, MA), levels 1, 2 and 3. In this example, the normal QC spectrum was obtained by averaging the sample spectra QC in the three lots 1 -3 in each level. Therefore, the deviations observed in this example arise from the differences in the sample lots QC. However, deviations from the normal spectrum are representative of deviations that could be observed from instrumental errors. Absorbance spectra were obtained in a CARY IV (Varian Instruments, Palo Alto, CA) and are presented in Figure 1 for QC level 1 lots 1 -3. The error spectra were set at seven wavelengths on the 550-650 nm scale, using absorbance spectra of H H b, O 2 Hb, COHb, MetHb, S H b, and lipid. The results are presented later. As can be seen, the values of the concentrations of the blood components, calculated using the method herein, correspond to values that can be observed for real blood. In contrast, the values of the blood component concentrations using the methods of the prior art can be negative or greater than 100% of the total hemoglobin.

LEVEL 1 QC-Tnom = 1 1 .5 gm / dL Average Concentration of QC Blood Component Concentration Errors Reported Results (Fnom,? X 100%) LEVEL QC 2 - Tnom = 17.5 gm / dL Average Concentration of QC Blood Component Concentration Errors Reported Results (Fnom i x 100%) LEVEL QC 3 - Tnom = 20.8 gm / dL Average Concentration of QC Blood Component Concentration Errors Reported Results (Fn0m, i x 100%) EXAMPLE 2 The QC level 1 spectra, lots 1 -3 presented in Figure 1, were fixed using a least squares analysis through the blood component spectra of Figure 2. The difference between the best fixation and the spectra QC is presented in Figure 3. As can be seen, the differences are orders of magnitude greater than the error spectrum obtained by subtracting the QC spectra of better fixation of the measured QC spectrum.

Claims (15)

1 .- A method to determine the performance of an absorbance spectrometer, the method includes: a. measure the absorbance spectrum of a quality control sample; b. fix the sample spectrum measured with the normal spectrum of the quality control product sample to obtain a spectrum of better fixation and an apparent concentration of the quality control sample; c. evaluate the quality of the fixation.

2. The method according to claim 1, wherein the quality of the fixation is evaluated by calculating the difference between the spectrum of best fixation and the spectrum measured to produce an error spectrum.

3. The method according to claim 1, wherein the quality of the fixation is evaluated by determining the deviation of the apparent concentration of the quality control sample, capp, from the concentration of the normal quality control sample, cstd-

4. The method according to claim 2, wherein a least squares analysis is used to set the normal spectrum to the measured spectrum.

5. The method according to claim 3, wherein a least squares analysis is used to set the normal spectrum to the measured spectrum.

6. The method according to claim 1, wherein the quality of the binding is evaluated by setting one or more blood component spectra to the error spectrum to produce errors in blood component concentrations.

7. The method according to claim 6, wherein a least squares analysis is used to set the normal spectrum for quality control to the measured spectrum to set one or more of the blood component spectra to the error spectrum .

8. The method according to claim 1, wherein the quality of fixation is evaluated by selecting a nominal total concentration of hemoglobin, multiplying the nominal total concentration of hemoglobin by the concentration of apparent quality control sample to produce a total concentration apparent hemoglobin

9. The method according to claim 8, wherein a least squares analysis is used to set the normal spectrum for the quality control sample to the measured spectrum.

10. The method according to claim 6, wherein the quality of the fixation is evaluated by selecting nominal values of concentrations of the blood components, adding the errors in concentrations of blood components to their respective nominal concentration values, to produce apparent concentrations of blood components, and evaluating the apparent concentrations of blood components.

11. - The method according to claim 10, wherein a least squares analysis is used to set the normal spectrum, for the quality control sample, to the measured spectrum and to set one or more of the blood component spectra to the spectrum of mistake.

12. The method according to claim 10, wherein the quality of the binding is determined by selecting a nominal total concentration of hemoglobin, multiplying the nominal total concentration of the hemoglobin by the concentration of the apparent quality control sample to produce a apparent total concentration of hemoglobin, and dividing the nominal concentrations of hemoglobin components by the apparent total concentration of hemoglobin to obtain fractional concentrations of hemoglobin components.

13. The method according to claim 6, wherein the concentration of the blood components are weighted to reflect anticipated instrument variations.

14. The method according to claim 1, wherein the normal quality control product has similar connotations of one or more absorption components of the quality control sample.

15. The method according to claim 1, further comprising the step of measuring the absorbance spectrum of a test sample. 16 - A method for carrying out an absorbance spectrophotometer analysis comprising: a. Measure the absorbance spectrum of a quality control product; b. setting the spectrum measured with a normal spectrum of a quality control product having the similar concentration of one or more absorbent components to obtain a spectrum of better fixation and an apparent concentration of the quality control sample; c. evaluate the quality of fixation; and d. Measure the absorbance spectrum of a test sample. 17. A method for calibrating an absorbance spectrophotometer comprising: a. measuring the absorbance spectrum for a plurality of quality control samples, wherein said quality control samples correspond to a spectrum of blood component; b. fix the spectrum measured with a normal spectrum of a quality control product that has the same concentration of absorbent components to obtain the spectrum of better fixation and an apparent concentration of the quality control sample; c. evaluate the quality of fixation; and d. adjust the caliber of said spectrophotometer.