Medical Device Link.

Originally Published IVD Technology March 2004

Assay Development

Ischemia and Stroke

Strokes are either occlusive (caused by blockage of a blood vessel owing to clot or other obstruction) or hemorrhagic (caused by bleeding from a vessel). Symptoms of stroke include sudden numbness or weakness, especially on one side of the body; sudden confusion or difficulty in speaking or understanding speech; sudden trouble seeing with one or both eyes; sudden difficulty in walking; dizziness; and loss of balance or coordination. Occlusion of oxygen-rich blood flow to the brain leads to localized tissue anemia, or ischemia, and damage to the nervous system.

TIA

A transient ischemic attack (TIA) is a short-lived episode of focal neurologic deficit that often precedes cerebral infarction. People who have suffered a TIA have 9.5 times greater risk of having a future stroke than those who have not had a TIA. After a first TIA, 10–20% of patients will likely suffer a stroke within the next 90 days; in 50% of these patients, the stroke occurs within the first 2 days. Most symptoms of a TIA disappear within an hour, although they may persist for up to 24 hours. Because symptoms are short-lived, many victims do not recognize the event as a TIA or do not perceive it to be a warning of a potentially impending stroke. This unawareness precludes the opportunity to initiate treatment in order to forestall possible onset of brain infarction.

The Processes of Ischemia

During the early stages of ischemia, cells are deprived of oxygen, triggering embolic and thrombotic processes (see Figure 2 in text for a schematic illustration of the neurotoxic cascade accompanying ischemia). Energy deficit occurs. Under oxidative stress, glucose undergoes transformation through pyruvate and a-keto-glutarate to glutamate, leading to lactate acidosis. Lack of energy causes the gated channels of the cell membrane that normally maintain homeostasis to open, allowing toxic amounts of calcium, sodium, and potassium ions to flow into the cell. The toxicity causes swelling and changes in intracellular osmotic pressure and membrane permeability.

Concurrently, injured ischemic cells release excitatory amino acids, glutamate, aspartate, and homocysteine, thus reducing membrane potentials. The immune system, meanwhile, is causing brain injury through an inflammatory reaction mediated by the vascular system. Microglia and astrocytes initiate local immunocompetent reactions and neuropeptide and antibody production. The further loss of membrane integrity results in damaged neurons; interaction of glial and microglial cells with vessel or capillary endothelial surfaces results in disruption of the blood-brain barrier.

An excessive amount of excitatory amino acids overactivates glutamate receptors, particularly N-methyl-D-aspartate (NMDA) receptors, which contribute to the pathological process of ischemic injury. Overstimulation of glutamate receptors allows excessive Ca++ influx into the cell, resulting in activation of various enzymes and proteases that begin to destroy glutamate receptors on neuronal membranes. Since the blood-brain barrier is compromised during ischemia, neuronal-membrane protein fragments resulting from the deterioration of cell membranes may enter the bloodstream at increased levels and initiate a general immune response. (The blood-brain barrier prevents the immune system from encountering these proteins under normal conditions.)

Stroke and Brain Damage

All of these processes culminate in stroke onset, with the symptoms manifested depending on whether cortical brain areas or deep structures were damaged.

The intensity of the neurotoxic cascade determines the death or survival of neuronal cells and the development of the ischemic penumbra, a territory surrounding the damaged brain area in which cells remain at risk for death. During this stage, administration of neuroprotective therapy within three hours of stroke onset is directed toward the recovering penumbra. Such prompt treatment is crucial for satisfactory patient outcome and prognosis.

To determine the accuracy of the PDDLS/Batch molecular size analysis system, a polystyrene standard having a nominal particle size diameter of 90 nm (and thus a radius of 45 nm) was diluted as an aqueous suspension and introduced to the detector.

The correlator was set for a sample time of 10 microseconds, a run time of 3 seconds, and a smoothing factor of 10. The last channel selected was channel number 1024. The assay time was only 1 minute. The sample distribution was monodisperse with a resultant Rh of 44.8 nm, as shown in Figure 2.

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