According to today's research, botulinum toxins are the strongest identified group of natural toxins, and they are produced by species of bacteria of the genus Clostridium that cause the development of a disease called botulism with characteristic acute symmetrical paralysis. Due to the severity of the symptoms caused by these bacteria in humans, with a possible fatal outcome for humans, any prevention of botulism disease is considered important, which requires the development of quality methods for determining botulinum toxin. The purpose of the work is to analyse the existing methods for determining botulinum toxin, whereby they are divided into in vivo and in vitro methods. It is considered that in vivo methods, although they ensure great precision, require longer periods of time for determination, which for certain types of toxins amount up to a couple of days. In addition to the above, in vivo methods are more expensive than in vitro methods due to the use of a larger number of living laboratory animals. In contrast to them, in vitro methods shorten the time required for analysis, but often show insufficient sensitivity. With the advancement of technology, the problems of accuracy and efficiency of methods are increasingly being solved, and scientists are successfully developing in vitro determination methods with great sensitivity and precision. Some of the best in vitro methods are immunoassays, analyses based on biosensors and fluorescence analyses.