The invention relates to the field of biological detection, in particular to a helicobacter pylori antigen colloidal gold detection kit, and a preparation method and an application thereof. The helicobacter pylori antigen colloidal gold detection kit provided by the invention comprises a test paper card, wherein the test paper card comprises a bottom plate, a sample pad, a gold mark pad, a nitrocellulose membrane and an absorbent pad; the sample pad, the gold mark pad, the nitrocellulose membrane and the absorbent pad are located on the surface of the bottom plate and are sequentially arranged from a sampling end; the gold mark pad comprises a colloidal gold-marked anti-helicobacter pylori monoclonal antibody-1; and the nitrocellulose membrane is coated with a detection line and a quality control line. The helicobacter pylori antigen colloidal gold detection kit provided by the invention has sensitivity and specificity and has the advantages of being fast, simple and convenient to operate, accurate in result, economical and practical.