APTT

APTT remains an option for monitoring UFH. It is very simple, quick and cheap project. However, it is difficult to standardise. The APTT assay requires that all proteins in the entire coagulation waterfall be intact, allowing accurate measurement of heparin levels. Patients with lupus anticoagulants or antiphospholipid syndrome often have elevated APTT, and they must be monitored with heparin tests. In addition to factors that vary in clotting factor levels, reagents and instruments can affect their sensitivity to heparin, resulting in results that can ultimately vary as much as fourfold between laboratories. The International Society for Thrombosis and HemoStasis and Standardization Committee (ISTH SSC) has attempted to establish a standard method for establishing a correction coefficient for APTT trials, similar to the ISI used in PT trials. But because of the variety of phospholipids, activators and instruments, this work has made only modest progress. ISTH recommends that each APTT assay system be calibrated by heparin anti-XA activity to obtain the appropriate APTT monitoring range using the recommended method.

Protamine sulfate neutralization test

The test is based on the principle that UFH, a highly negatively charged molecule, is neutralized by protamine sulfate, a positively charged protein. Protamine sulfate with different concentrations was added to the plasma, and then thrombin was added to measure the coagulation time. The concentration of protamine sulfate that restores thrombin coagulation time to normal is considered to be the concentration of heparin, and this process is only used at UFH.

Assay of anti-Xa activity

The principle of detecting the anti-XA activity of heparin by the chromogenic substrate method is the same: heparin in the specimen forms a complex with AT to inhibit the excessive addition of factor Xa. The activity of the remaining factor Xa was measured by its interaction with specific substrates to release pNA. This reaction is inversely proportional to the concentration of heparin. The only differences are the incubation time of the reaction, the buffer used for dilution, the substrate, and whether or not exogenous AT is added. Glucan sulfate in the buffer can reduce the effect of PF4.

One step detection

The one-step method of measuring heparin makes the test simpler and shortens the time. This method does not add an exogenous AT.

Detection is based on the principle of competition suppression. Xa was added to the plasma to mix with the substrate and immediately there were 2 simultaneous reactions: hydrolysis of the substrate by Xa and inhibition of Xa by the heparin-AT complex. Once the reaction is in equilibrium, the pNA released from the substrate is inversely proportional to the concentration of heparin in the plasma under test. In this test, no exogenous AT is added and it depends entirely on the amount of AT in the patient's plasma. This represents the patient's true heparin functional response. The patient's AT level was between 35% and 130%, and the heparin concentration test was not affected.