The knowledge on spiders of the Czech Republic has substantially improved since the second version of the national Red List was published, mainly due to large-scale field records and the establishment of an extensive, searchable electronic database of both retrospective and prospective records. Meanwhile, Central European spiders have undergone substantial changes in abundance and distribution. In this report, an updated Red List is presented and compared with the previous editions from 1992 and 2002. Here, we assessed all 879 spider species known to occur in the Czech Republic. For the first time, the abundance, area of occupancy and population trends were calculated for each of the species using the data from the Czech Arachnological Society recording scheme. Twenty-seven species (3% of the total) were classified as Regionally Extinct (RE), 94 (11%) as Critically Endangered, 113 (13%) as Endangered, 156 (18%) as Vulnerable, and 126 (14%) were classified as Least Concern species. Some species listed in the previous version of the Red List were found to live also in non-endangered habitats or to be more common than previously thought,and were thus removed from the list or reclassified to the lower Red List categories. Additionally, several species with dramatically decreasing abundance were identified, among them Ozyptila rauda, Agyneta equestris, Agyneta mollis, Kishidaia conspicua, Clubiona genevensis and Centromerus semiater. The results confirm that spiders are a highly threatened group of arthropods in the Czech Republic, and the updated Red List provides an important foundation for defining conservation priorities.