Methods to determine a diagnostic indicator of blood sugar conditions, and, liquid chromatographic columns therefor (cyanide free)

Claim 1

A method to determine a numerical percentage value as a diagnostic indicator of the blood sugar condition of a specific person, wherein a whole blood sample is taken from said person and thereafter prepared as a test sample containing a red blood cell hemolysate solution, and thereafter, a quantity of said test sample is introduced into an end of a column bed having no cyanide therein which will adsorb hemoglobin species present in said test sample, said column bed comprising an equilibrated suspension of ion exchange material particles having a size less than 100 mesh, said particles in a column bed being one member selected from the class consisting of CarXH and CarYOH, where "Car" represents an inert substrate for carrying ionizable groups X 31 providing dissociated cations H + and ionizable groups Y + providing dissociated anions OH - , said CarXH particles being a weakly acidic cation exchanger having a reported pK a : 3-7 and being used in an equilibrated suspension at a pH: 6.0-7.5 at 22.5° C., said CarYOH particles being a weakly basic anion exchanger having a reported pK a : 7-10 and being used in an equilibrated suspension at a pH: 7.3-9.0 at 22.5° C., and thereafter, a quantity of a buffer solution having no cyanide therein is introduced into an end of said column bed to preferentially elute therefrom a first fraction which contains certain of the hemoglobin species present in said test sample, a column bed which is an equilibrated suspension of CarXH particles providing a said first fraction containing substantially all of the hemoglobin species Hb-A 1a-c present in said test sample, a column bed which is an equilibrated suspension of CarYOH particles providing a said first fraction containing essentially all of the hemoglobin species present in said test sample other than Hb-A 1a-c , and thereafter, an aliquot quantity of said first eluate fraction is collected from the other end of said column bed, and then, a quantity of a wash solution is introduced into an end of said column bed to desorb and elute therefrom a second fraction containing substantially all of the reminaing hemoglobin species present in said test sample, a column bed which is an equilibrated suspension of CarXH particles providing a said second fraction containing essentially all of the hemoglobin species present in said test sample other than Hb-A 1a-c , a column bed which is an equilibrated suspension of CarYOH particles providing a said second fraction containing substantially all of the hemoglobin species Hb-A 1a-c present in said test sample, and then, an aliquot quantity of said second eluate fraction is collected from the other end of said column bed, and then, the hemoglobin species present in the said first and second eluate fractions are separately detected and measured by spectrometric analysis and the respective amounts thereof are expressed as numerical values which are then compared in accordance with a mathematical formula to provide a numerical percentage value for the hemoglobin species Hb-A 1a-c in said test sample for use as a diagnostic indicator of the blood sugar characteristics of said specific person.

Claim 2

A method to determine a numerical percentage value as a diagnostic indicator of the blood sugar condition of a specific person, wherein a whole blood sample is taken from said person and thereafter prepared as a test sample containing a red blood cell hemolysate solution, and thereafter, a quantity of said test sample is introduced into an end of a column bed having no cyanide therein which will adsorb hemoglobin species present in said test sample, said column bed comprising an equilibrated suspension of ion exchange material particles having a size less than 100 mesh, said particles in a column bed being one member selected from the class consisting of CarXH and CarYOH, where "Car" represents an inert substrate for carrying ionizable groups X 31 providing dissociated cations H + and ionizable groups Y + providing dissociated anions OH - , said CarXH particles being a weakly acidic cation exchanger having a reported pK a : 3-7 and being used in an equilibrated suspension at a pH: 6.0-7.5 at 22.5° C., said CarYOH particles being a weakly basic anion exchanger having a reported pK a : 7-10 and being used in an equilibrated suspension at a pH: 7.3-9.0 at 22.5° C., and thereafter, a quantity of a buffer solution having no cyanide therein is introduced into an end of said column bed to preferentially elute therefrom a first fraction which contains certain of the hemoglobin species present in said test sample, a column bed which is an equilibrated suspension of CarXH particles providing a said first fraction containing substantially all of the hemoglobin species Hb-A 1a-c present in said test sample, a column bed which is an equilibrated suspension of CarYOH particles providing a said first fraction containing essentially all of the hemoglobin species present in said test sample other than Hb-A 1a-c , and thereafter, an aliquot quantity of said first eluate fraction is collected from the other end of said column bed, and then, a quantity of a wash solution is introduced into an end of said column bed to desorb and elute therefrom a second fraction containing substantially all of the reminaing hemoglobin species present in said test sample, a column bed which is an equilibrated suspension of CarXH particles providing a said second fraction containing essentially all of the hemoglobin species present in said test sample other than Hb-A 1a-c , a column bed which is an equilibrated suspension of CarYOH particles providing a said second fraction containing substantially all of the hemoglobin species Hb-A 1a-c present in said test sample, and then, an aliquot quantity of said second eluate fraction is collected from the other end of said column bed, and then, the hemoglobin species present in the said first and second eluate fractions are separately detected and measured by spectrometric analysis and the respective amounts thereof are expressed as numerical values which are then compared in accordance with a mathematical formula to provide a numerical percentage value for the hemoglobin species Hb-A 1a-c in said test sample for use as a diagnostic indicator of the blood sugar characteristics of said specific person.

Claim 3

A method to determine a numerical percentage value as a diagnostic indicator of the blood sugar condition of a specific person, wherein a whole blood sample is taken from said person and thereafter prepared as a test sample containing a red blood cell hemolysate solution, and thereafter, a quantity of said test sample is introduced into an end of a column bed having no cyanide therein which will adsorb hemoglobin species present in said test sample, said column bed comprising an equilibrated suspension of ion exchange material particles having a size less than 100 mesh, said particles in a column bed being one member selected from the class consisting of CarXH and CarYOH, where "Car" represents an inert substrate for carrying ionizable groups X - providing dissociated cations H + and ionizable groups Y + providing dissociated anions OH - , said CarXH particles being a weakly acidic cation exchanger having a reported pK a : 3-7 and being used in an equilibrated suspension at a pH: 6.0-7.5 at 22.5° C., said CarYOH particles being a weakly basic anion exchanger having a reported pK a : 7-10 and being used in an equilibrated suspension at a pH: 7.3-9.0 at 22.5° C., and thereafter, a quantity of a buffer solution having no cyanide therein is introduced into an end of said column bed to preferentially elute therefrom a first fraction which contains certain of the hemoglobin species present in said test sample, a column bed which is an equilibrated suspension of CarXH particles providing a said first fraction containing substantially all of the hemoglobin species Hb-A 1a-c present in said test sample, a column bed which is an equilibrated suspension of CarYOH particles providing a said first fraction containing essentially all of the hemoglobin species present in said test sample other than Hb-A 1a-c and thereafter, an aliquot quantity of said first eluate fraction is collected from the other end of said column bed, and then, a quantity of said test solution is significantly diluted to provide a red blood cell hemolysate solution which may be conveniently detected by spectrometric analysis, and then, the hemoglobin species present in said first eluate fraction and in the said diluted hemolysate solution are separately detected and measured by spectrometric analysis and the respective amounts thereof are expressed as numerical values which are then compared in accordance with a mathematical formula to provide a numerical percentage value for the hemoglobin species Hb-A 1a-c in said test sample for use as a diagnostic indicator of the blood sugar characteristics of said specific person.

Claim 4

A method to determine a numerical percentage value as a diagnostic indicator of the blood sugar condition of a specific person, wherein a whole blood sample is taken from said person and thereafter prepared as a test sample containing a red blood cell hemolysate solution, and thereafter, a quantity of said test sample is introduced into an end of a column bed having no cyanide therein which will adsorb hemoglobin species present in said test sample, said column bed comprising an equilibrated suspension of ion exchange material particles having a size less than 100 mesh, said particles in a column bed being one member selected from the class consisting of CarXH and CarYOH, where "Car" represents an inert substrate for carrying ionizable groups X - providing dissociated cations H + and ionizable groups Y + providing dissociated anions OH - , said CarXH particles being a weakly acidic cation exchanger having a reported pK a : 3-7 and being used in an equilibrated suspension at a pH: 6.0-7.5 at 22.5° C., said CarYOH particles being a weakly basic anion exchanger having a reported pK a : 7-10 and being used in an equilibrated suspension at a pH: 7.3-9.0 at 22.5° C., and thereafter, a quantity of a buffer solution having no cyanide therein is introduced into an end of said column bed to preferentially elute therefrom a first fraction which contains certain of the hemoglobin species present in said test sample, a column bed which is an equilibrated suspension of CarXH particles providing a said first fraction containing substantially all of the hemoglobin species Hb-A 1a-c present in said test sample, a column bed which is an equilibrated suspension of CarYOH particles providing a said first fraction containing essentially all of the hemoglobin species present in said test sample other than Hb-A 1a-c and thereafter, an aliquot quantity of said first eluate fraction is collected from the other end of said column bed, and then, a quantity of said test solution is significantly diluted to provide a red blood cell hemolysate solution which may be conveniently detected by spectrometric analysis, and then, the hemoglobin species present in said first eluate fraction and in the said diluted hemolysate solution are separately detected and measured by spectrometric analysis and the respective amounts thereof are expressed as numerical values which are then compared in accordance with a mathematical formula to provide a numerical percentage value for the hemoglobin species Hb-A 1a-c in said test sample for use as a diagnostic indicator of the blood sugar characteristics of said specific person.