Claims Analysis
Total Claims: 15
Claim 1
Independent Claim
151 words
An in vitro method for effecting rapid separation of the MM and MB creatine kinase isoenzymes in blood plasma or serum and for determining the relative activities of the thus separated isoenzymes comprising the steps of contacting a blood plasma or serum sample and a buffer of low ionic strength with an ion exchange support comprising porous glass beads having a stationary phase coupled to the surfaces thereof through an intermediate coupling agent; incubating the resultant mixture to effect MB isoenzyme adsorption by said support; separating the supernatant liquid fraction containing the MM isoenzyme from said support; washing the support to effect removal of residual MM; adding a buffered solution of a strong electrolyte to said support to effect desorption of the MB isoenzyme therefrom; separating the supernatant liquid fraction containing the MB isoenzyme from said support; and assaying the MM and MB activity in the respective liquid fractions thus obtained.
Claim 2
Dependent Claim
151 words
An in vitro method for effecting rapid separation of the MM and MB creatine kinase isoenzymes in blood plasma or serum and for determining the relative activities of the thus separated isoenzymes comprising the steps of contacting a blood plasma or serum sample and a buffer of low ionic strength with an ion exchange support comprising porous glass beads having a stationary phase coupled to the surfaces thereof through an intermediate coupling agent; incubating the resultant mixture to effect MB isoenzyme adsorption by said support; separating the supernatant liquid fraction containing the MM isoenzyme from said support; washing the support to effect removal of residual MM; adding a buffered solution of a strong electrolyte to said support to effect desorption of the MB isoenzyme therefrom; separating the supernatant liquid fraction containing the MB isoenzyme from said support; and assaying the MM and MB activity in the respective liquid fractions thus obtained.
Claim 3
Dependent Claim
51 words
An in vitro method as set forth in claim 1 wherein said intermediate coupling agent is a silane coupling agent constituted by an organosilane with a silicon functional group capable of bonding to the surface of the glass beads and an organic functional group capable of bonding to the stationary phase.
Claim 4
Dependent Claim
22 words
An in vitro method as set forth in claim 2 wherein said organic functional group is a simple carbohydrate or carbohydrate derivative.
Claim 5
Dependent Claim
17 words
An in vtro method as set forth in claim 3 wherein said organic functional group is glycerol.
Claim 6
Dependent Claim
18 words
An in vitro method as set forth in claim 2 wherein said intermediate organosilane coupling agent is glycidoxypropyltrimethoxysilane.
Claim 7
Dependent Claim
20 words
An in vitro method as set forth in claim 2 wherein said stationary phase is derived from diethylamine or diethylaminoethanol.
Claim 8
Dependent Claim
34 words
An in vitro method as set forth in claim 1 wherein said intermediate coupling agent is formed from a Grignard reagent of the formula RMgX where R is lower alkenyl and X is halogen.
Claim 9
Dependent Claim
18 words
An in vitro method as set forth in claim 7 wherein said Grignard reagent is 1-butenyl magnesium bromide.
Claim 10
Dependent Claim
25 words
An in vitro method as set forth in claim 1 wherein the buffer of low ionic strength has a pH between approximately 7.1 and 9.0.
Claim 11
Dependent Claim
27 words
An in vitro method as set forth in claim 1 wherein the concentration of strong electrolyte in said buffered solution is between approximately 250 and 500 mmol/liter.
Claim 12
Dependent Claim
16 words
An in vitro method as set forth in claim 10 wherein the electrolyte is sodium chloride.
Claim 13
Dependent Claim
47 words
An in vitro method as set forth in claim 1 wherein the sample is blood plasma and the step of washing the support to effect removal of residual MM results in at least approximately a 1000 fold dilution of MM activity in the initial blood plasma sample.
Claim 14
Dependent Claim
24 words
An in vitro method as set forth in claim 1 wherein the resultant mixture from the first step is incubated for approximately three minutes.
Claim 15
Dependent Claim
25 words
An in vitro method as set forth in claim 1 wherein said incubating and separating steps are carried out by means of a filter sampler.