In order to analyze formaldehyde, the simplest aldehyde closely related to human health from both exogenous and endogenous surroundings, many methods have been developed and employed with different techniques and instruments. In fact, measurements by different methods result in different values for the concentration of exogenous and endogenous formaldehyde. Among the methods, high-performance liquid chromatography (HPLC) is one of the most commonly used approaches in the determination of formaldehyde in the liquid and body tissues such as urine, blood, the brain, and liver. HPLC and gas chromatography (GC) coupled with mass spectrometry (MS) or fluorescent techniques show their high sensitivity and advantages. 2,4-Dinitrophenylhydrazine (DNPH) is one of the most frequently used reagents applied with spectrophotometer, HPLC, and GC. HPLC coupled with DNPH or ampicillin has been used to determine formaldehyde levels of the morning urine from Alzheimer’s disease (AD) patients and age-matched participants. The results showed a positive correlation between the levels of urine formaldehyde and the severity of cognitive impairment in the AD patients. Approximately 40% of the AD patients have got high levels of endogenous formaldehyde, suggesting that endogenous formaldehyde can be developed as a biomarker for progression and intervention of AD. Currently, the specific fluorescent probes such as formaldehyde probe-1 (FAP-1) and formaldehyde-induced 2-aza-Cope reaction have been synthesized to label formaldehyde molecules in cells. In particular of using these fluorescent probes, the endogenous formaldehyde is found to be localized in organelles such as lysosomes and mitochondrion. Exogenous formaldehyde is also localized and restricted in lysosome inside a living cell monitored by confocal microscopy. Furthermore, nano-techniques have been also introduced to analyze formaldehyde. Quartz crystal microbalance (QCM) can measure nanogram-scale changes of formaldehyde. As the technique advances, current efforts are underway to develop greater sensitivity, lower cost, improved portability, and more straightforward detection methods. The operational principles and sensing performance of sensors have been created as well, and the convenient and high-performance formaldehyde-determining applications will appear more in the near future.