A research group led by Assistant Professor Mitsuru Hattori and Professor Takeharu Nagai of the Institute of Scientific and Industrial Research, Osaka University has developed a bioluminescence indicator called Thrombastor that can rapidly measure the activity of thrombin, an enzyme that promotes blood coagulation.
Thrombotic conditions such as cerebral infarction and myocardial infarction are caused by the process of blood coagulation in blood vessels and formation of thrombi. Thrombin is an enzyme that plays a central role in blood coagulation, and measuring its activity leads to the prediction of thrombosis and prevention of its onset.
In this research, the research group developed a new indicator, Thrombastor, that can measure thrombin activity in blood using bioluminescence, which is the same process by which fireflies emit light. Normally, Thrombastor emits green light; however, this light turns blue when Thrombastor is cleaved by thrombin. Thrombin activity can be measured by quantifying changes in emission color. Additionally, because Thrombastor very brightly emits light, it is possible to capture photos using general-purpose cameras such as a smartphone camera. To verify that thrombin activity in the blood is actually measurable, researchers attempted detection using Thrombastor.
Thrombastor is composed a bioluminescent protein, NanoLuc and a fluorescent protein, Gamillus linked by thrombin recognition sequence. When the sequence is cleaved by thrombin, resonance energy transfer is cancelled, resulting in a change in the emission color from green to blue.
Provided by Osaka University
Thrombastor was added to plasma isolated from mouse blood, and the luminescence was photographed using a smartphone camera. As a result, the emission color was observed to change from green to blue depending on the time from the collection to the end of the coagulation reaction. This color change confirmed that Thrombastor was cleaved by thrombin produced in plasma over time.
After mixing several μL of collected blood plasma with Thrombastor, the bioluminescence was generated by the addition of substrate. This system measures the activity of thrombin produced in the blood by analyzing images taken with a smartphone camera at the ratio of green to blue.
Provided by Osaka University
"By using a smartphone to take a picture of the emission color change (from green to blue) of the indicator according to the thrombin activity, you can easily measure thrombin activity in the blood. If this method can be developed into a simple examination method that can be used at home, we believe it will function as a technology that plays a part in home medical care," says Professor Nagai.
■ Thrombin: An enzyme (protease) involved in blood coagulation. It catalyzes the hydrolysis of fibrinogen in the blood to form insoluble fibrin. It is present as prothrombin in plasma and is activated by factor V during bleeding to form thrombin.
This article has been translated by JST with permission from The Science News Ltd.(https://sci-news.co.jp/). Unauthorized reproduction of the article and photographs is prohibited.