Chemistry: analytical and immunological testing – Involving an insoluble carrier for immobilizing immunochemicals
Reexamination Certificate
2000-11-15
2003-07-01
Le, Long V. (Department: 1641)
Chemistry: analytical and immunological testing
Involving an insoluble carrier for immobilizing immunochemicals
C436S523000, C436S524000, C436S525000, C436S526000, C436S823000, C435S007100, C435S007210, C435S007920, C435S007240
Reexamination Certificate
active
06586259
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to a platelet/leukocyte interaction assay allowing for point of care assessment of interaction of platelets and leukocytes, and the reagents therefor.
2. Discussion of the Background
Platelets are known to interact with leukocytes both as a consequence of contact during normal blood flow (Stone and Nash,
British Journal of Haematology
, 105:514-22, 1999; Lorenz et al,
Blood Coagulation and Fibrinolysis
, 9:S49-59, 1998) and as a consequence of various pathological processes (Rinder et al,
Journal of Cardiovascular Surgery
, 118:460-6, 1999; Pevton et al,
Journal of Vascular Surgery
, 27:1109-15, 1998; Stuard et al,
International Journal of Artificial Organs
, 21:75-82, 1998; Gawaz et al,
European Journal of Clinical Investigation
, 25:843-51, 1995).
Pathological conditions such as Unstable Angina, Coronary Artery Disease (CAD), and Stroke are characterized by high levels of platelet and leukocyte activity. Measurement of platelet/leukocyte interaction can be predictive of these pathological states, particularly in combination with other diagnostic factors. Also, measurement of platelet/leukocyte interaction can be used as a means of monitoring therapy directed toward altering platelet and/or leukocyte function.
Exposure of flowing blood to artificial surfaces has been shown to enhance platelet/leukocyte interaction. The cell types involved and the extent of the interaction vary with the composition of the artificial surface in contact with the blood (Gawaz et al,
Artificial Organs
, 23:29-36, 1999).
Although platelet/leukocyte interactions have been quantified using various techniques (Hendricks et al, U.S. Pat. No. 5,503,982; Rinder et al,
Blood
, 78:1760, 1991;), assessment of the interaction has relied upon measurement of circulating platelet/leukocyte complexes. Measurements, to date, have taken the form of evaluating pre-existing platelet/leukocyte interactions in a blood sample.
In some pathological conditions (e.g. Acute Myocardial Infarction, AMI; post Angioplasty, PTCA; etc.) platelet/leukocyte complex formation is associated with interaction with damaged subendothelium, whether directly (plaque formation) or indirectly (release of biochemical markers such as ICAM-1, see Hendricks et al, U.S. Pat. No. 5,503,982).
Current assay systems used to assess platelet/leukocyte interactions, as exemplified by Hendricks et al (U.S. Pat. No. 5,503,982), evaluate pre-existing (circulating) platelet/leukocyte complexes and do not utilize a component representative of the vessel subendothelium (i.e. extracellular matrix) or other solid-phase stimulus. Moreover, the threshold at which discrete platelets and leukocytes interact could vary depending on the activation status of these cells at the time of testing. It is known in the art that platelet and/or leukocyte activation is a necessary prerequisite of platelet/leukocyte binding. It is also known in the art that certain pathological conditions are associated with upregulation of platelet and/or leukocyte activity. However, the upregulation in cellular activity associated with the pathological process may be insufficient to support platelet/leukocyte complex formation without additional stimulation and moreover, may not be detectable using conventional systems, which lack a stabilizing solid-phase support upon which the platelet/leukocyte complex could be maintained. (A solid-phase stimulus could be used as a means of localizing pre-existing platelet leukocyte complexes and/or inducing complex formation and localization in cells predisposed to do so.)
It is desirable that assay systems designed to incorporate the use of a solid-phase component such as immobilized subendothelial/extracellular matrix be facile, rapid and of reasonable cost to be useful in detecting platelet/leukocyte interaction in a clinical setting.
The present invention addresses shortcomings of previous methods and technologies by using microparticles of various compositions coated with plasma proteins and/or extracellular matrix proteins, either singly or in combination, to facilitate rapid assessment of platelet/leukocyte binding.
Platelets can interact with leukocytes through various mechanisms, such as contact during normal blood flow (Lorenz et al,
Blood Coagulation and Fibrinolysis
, 9:S49-S59, 1998), or as a consequence of a pathological process associated with platelet hyperactivity (Spanygenberg,
Thrombosis Research
, 74:S35-S44, 1994; Rinder et al,
Journal of Cardiovascular Surgery
, 118:460-6, 1999) or due to an inflammatory process (Gawaz et al,
European Journal of Clinical Investigation
, 25:843-51, 1995). Receptors found on the platelet surface interact with receptors found on various leukocytes through direct bridging or through an indirect linkage involving intermediary molecules (Weber and Springer,
Journal of Clinical Investigation
, 100:2085-93, 1997). Upregulation of platelet and/or leukocyte activity favors enhanced platelet/leukocyte interaction (Rinder et al, 1999; Stone and Nash,
British Journal of Haematology
, 105:514-22, 1999; Konstantopoulos et al, 1998; Gawaz et al, 1995; Spanenberg, 1994).
Individuals with Coronary Artery Disease (CAD), Diabetes or Cerebrovascular Ischemia demonstrate both platelet hyperactivity and an ongoing inflammatory process (Michelson and Furman,
Current Opinion in Hematology
, 6:342-8, 1999). Treatment of patients with CAD has involved the use of anti-platelet agents and anti-inflammatory medications (Vorchheimer et al,
JAMA
281:1407-14, 1999; Mannaioni et al,
Inflammation Research
, 46:4-18, 1997).
Platelet/monocyte (Hendricks et al, U.S. Pat. No. 5,503,982) and platelet
eutrophil (Gawaz et al,
European Journal of Clinical Investigation
, 25:843-51, 1995) interaction have been suggested to be predictive of acute myocardial infarction (AMI) and inflammation, respectively. A corollary to platelet/leukocyte interaction in various pathological conditions is involvement of the vessel wall, whether in plaque formation for example, or a localized inflammatory reaction.
Platelet function assessment using immobilized extracellular matrix proteins has been described by Shaw and Stewart (U.S. Pat. No. 5,427,913). The authors demonstrated that von Willebrand factor (VWF) immobilized on polystyrene beads could be used to activate platelets and thereby determine the functional status of platelets from patients with platelet function defects. In addition, the authors also demonstrated that the effects of agents designed to alter platelet function could be monitored using bead-immobilized VWF as a stimulus. The results of these studies underline the importance of evaluating a normal hematological interaction or the hematologic consequence of a pathological state in the presence of an agent that mimics components of the vessel wall.
Although Shaw and Stewart describe methods and compositions of determining platelet function, there is no description or suggestion of using their method for evaluating platelet/leukocyte interaction.
CVDI's TAS™ analyzer measures the kinetics of fibrin polymerization following activation of the coagulation pathway in a patient's blood sample. The TAS™ analyzer and disposable were designed for use with whole blood in a point-of-care setting. Paramagnetic iron oxide particles (PIOP) are an essential component of the detection system for each of the tests developed for the TAS™ analyzer. The PIOP and other lyophilized ingredients for a particular test are located in the shallow reaction chamber of the TAS test card disposable. In addition to PIOP, the test reagent may contain buffers, stabilizers, fillers and specific coagulation pathway activator or agents. A test is initiated by insertion of a dry-chemistry test card into a slot of the TAS™ analyzer that automatically positions the test card reaction chamber above an electromagnet. This chamber is also illuminated with infrared light from a light emitting diode. The instrument measures reflected infrared light from the surface of the
Mahan Donald E.
Stewart Michael W.
Counts Gary
Le Long V.
Oblon & Spivak, McClelland, Maier & Neustadt P.C.
Pharmanetics Incorporated
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