Skeletal muscular myotubular dynamics measuring chip and preparation method thereof

Abstract

The invention relates to a skeletal muscular myotubular dynamics measuring chip and a preparation method thereof. The skeletal muscular myotubular dynamics measuring chip is suitable for measuring mechanical strength of skeletal muscular myotubes under contraction and is applicable to scientific researches in the biomedicine engineering field. The skeletal muscular myotubular dynamics measuring chip comprises a glass slide, a double-layer polymer substrate and a fluorescent label. The glass slide serves as a base for the chip and functions to support and stabilize the chip; the polymer substrate including two layers includes an upper layer and a lower layer that is thick, through different speeds of a spinner, the thickness of the lower layer is controlled 38 mu m and above, and the thickness of the upper layer is controlled to 10 mu m and below; the fluorescent label serves a core element to the chip, is arranged between the two layers of polymer material and is close to the upper surface, so that a displacement can be generated under the influence of contraction of surface skeletal myotubes. The skeletal muscular myotubular dynamics measuring chip and the preparation method thereof have the advantages that observation is easy, accuracy and visibility of myotubular contraction measurement can be greatly improved, operational difficulty is greatly reduced, myotubular cultivation, mechanical measurement and other experimental steps can be finished within a short period.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and designs a chip for measuring skeletal muscle myotube mechanics and a preparation method thereof, specifically a chip and a preparation method for visually and accurately observing and collecting mechanical information of skeletal muscle myotube contraction force. Background technique [0002] The need for techniques for measuring myotube mechanics emerged in this century. At present, the scientific research field is trying to establish a muscle tissue chip for physiological and pathological research and drug screening, providing a technical basis for muscle tissue diseases. At the beginning of the 21st century, researchers successfully constructed skeletal muscle tissue chips arranged in parallel through methods such as mechanics, electrical stimulation, and chemical modification. A large number of studies have shown that geometric arrangement will affect the differentiation efficiency of...

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Problems solved by technology

In 2000, Alenghat[1] et al. used magnetic tweezers to measure the contractility of single cells, but the cells need to endocytose the magnetic beads first. This method will seriously affect the normal life activities of cells.

In 2007, Wilson[2] et al. combined silicon cantilevers with atomic force microscopes to measure the contraction force of single-layer muscle fibers. However, the production process of silicon cantilevers is complicated, the operation is complicated, and the success rate is low. It is impossible to achieve a large number of repeated experiments, and the atomic force microscope is not very Common equipment, ordinary laboratories can not meet the corresponding conditions

However, these complex devices are very unrealistic for the high-throughput requirements of drug screening, and the success rate is also extremely low

In 2010, Mann[4] et al. used immunofluorescence microspheres to measure the contractility of myotubes, which can achieve more accurate mechanical measurements, but similarly, fluorescent microspheres will be endocytosed by cells, thereby affecting the life activities of the cells themselves , cannot objectively reflect the experimental results, and this method cannot control the arrangement and number of myotubes

In addition, this method cannot control the vertical height of fluorescent microspheres, which will also cause certain deviations in the later statistical results

In 2011, Fujita[5] and others used micro-electromechanical methods to measure, but this method still has low success rate, high equipment requirements, high cost, complicated operation, cannot achieve high-throughput requirements, and is not easy to control myotubes. Shape and quantity, myotubes will shrink to a ball and no longer open during operation

In 2013, the PDMS film cantilever technology used by Sun[6] can quantitatively measure the mechanical properties of muscle fibers, and can realize the quantitative determination of the number of muscle fibers, but this method needs to cut the PDMS film into millimeter-sized cantilever beams, release one end, The operation is complicated, and many cells and myotubes are destroyed during the operation, resulting in low success rate and low repeatability of the experiment

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