The key drawbacks of these methods are their pitiable sequential, spatial resolutions, and difficulty of the supplementary technical arrangements. Utilization of applicable micro-biosensors could overcome these difficulties with carbon-fiber-based electrodes looking generally efficient. For conventional methods for biochemical recognition including HPLC on micro-dialysis samples, it was used to immobilize BF-170 hydrochloride enzyme column and combined electrochemical detectors. A recent study was performed on pH sensitive poly Acepromazine maleate membrane with a plasma-polymerized film as a potentiometric biosensor for bio-chemical recognition, where the detection parameters were not satisfactory. However, for this plasmapolymerized film fabricated device, the characteristic curve was not linear, therefore calibration was urgently required. There are a lot of applications for Enzyme Field-Effect Transistors glucose, urea, acetylcholine, and alcohol using various enzymes. The technique of enzyme on biochips is very essential, where experimental immobilization background is concerned on sensitivity and stability. Biosensors based on the principle of field-effect in semiconductor structures have been comprehensively studied in recent years. Lately, it was potentially developed a charge-transfer-type pH sensor based on a charge-coupled devices, where sensitivity of devices was not achieved in satisfactory-level. With various methodology, the cholesterol biosensors were achieved a substantial interest owing to their sensitivity, selectivity, fast response time, repeatability, and stability. The mediator free electrochemical biosensors are based on suitable immobilization of selective enzyme on proper matrixes offers a portable, economical, disposable, and fast technique for the detection of different bio-molecules. In recent times, researchers are investigated with bio-compatible composite materials as appropriate matrixes for the enzyme immobilization for the efficient recognition of different biological molecules. Among various immobilization techniques, the Au/TGA/ChOx fabricated bio-chips are one of the most promising matrixes which can be used for the immobilization of enzymes due to their numerous interesting properties such as non-toxicity, mediator-less detection, high-surface area, requiring low-sample volume, fastresponse time, chemical stability, highly-sensitive, ease of handling, selective, and ease of enzymatic fabrication. In this report, it is developed highly sensitive mediator-free cholesterol biosensors based on tiny bio-chips at room conditions.