**Acknowledgements**

However, KKO is a mouse IgG2b antibody (an absent subclass in humans) [66], while the platelet-activating aPF4/P Abs present in HIT plasma samples are predominantly IgG1. KKO behaves differently from human aPF4/P Abs, i.e. it binds only weakly to PF4/H complexes coated on a solid phase [64]. Recently, a chimeric monoclonal aPF4/H Abs with a human Fc fragment (5B9) has been developed [67]. The 5B9 antibody has been demonstrated to fully

Immunologic assays, such as polytypic ELISA, IgG-specific ELISA, and particle gel immunoassay (PGI) have a sensitivity, are widely used to detect aPF4/H Abs in the diluted human sera because of their high sensitivity (≥95%) and the fast turn-around. However, only ~50% of aPF4/H Abs detected by antigen tests are clinically irrelevant. The results from positive immunologic assays may lead to an overtreatment for HIT that can result in serious consequences, such as venous limb gangrene or fatal hemorrhage [69]. However, immunologic assays are still powerful tools to rule-out patients with HIT. The cut-off optical intensity (OD) in ELISA was defined at 0.5. An ELISA test showing OD > 0.5 is normally suspected to contain aPF4/H Abs. To increase the specificity of clinically relevant antibodies, a higher OD cut-off

Even though functional assays such as by serotonin release assay (SRA) [71] or HIPA [72] have a sensitivity of ~90% which is slightly lower than the immunologic assays, these tests show a much better specificity of over 90%. For the better identifying HIT, it is recommended that a positive PF4/H ELISA should prompt confirmatory testing by functional assays [73]. However, the functional assays are only available in specialized laboratories and not available in many countries. Therefore, many physicians rely on the results of antigen tests, especially for the first days after clinical suspicion of HIT has been raised until the results of the functional assay is reported. Besides immunologic assays and functional assays, the chemiluminescent immunoassays such as HemosIL AcuStar HIT-IgG and HemosIL AcuStar HIT-Ab have been recently introduced. These methods are relatively faster (~30 minutes) than the immunologic assays (hours) and showed extremely high sensitivity (~100%) [74]. The assays seem to be ideal for ruling out HIT. Another study used a colorimetric test to detect HIT based on the interaction between platelets and tetrazolium-based indicator dye [75]. The authors reported the quality of detect-

Not only heparin but also autoimmune antibodies induce thrombocytopenia. Large antigenic complexes formed between PF4 and either heparin or antibody activate platelets, cause a prothrombotic and result in a variety of thromboembolic and systemic consequences. In autoimmune HIT, aPF4/P Abs activate platelets in the absence of heparin. These antibodies are highly reactive. They can self-cluster PF4-molecules forming antigenic complexes and allow

mimic the cellular effects of human HIT Abs [10, 68].

for the antigen tests (e.g. OD > 1.0) had been suggested [70].

ing HIT is from 96 to 100% agreement with the functional assay C-SRA.

**4. Diagnosis of HIT**

44 Thrombocytopenia

**5. Conclusion**

This work was supported by the Deutsche Forschungsgemeinschaft (DFG, Germany) (NG 133/1-1).
