The invention discloses a method and equipment for preparing an isosmotic disinfection cleaning solution. The method for preparing the isosmotic disinfection cleaning solution comprises the following steps of: performing ultrafiltration, active carbon filtering, micro-filtration and reverse osmosis treatment on raw water to obtain purified water; mixing the purified water with an NaCl aqueous solution to obtain electrolyte, feeding the electrolyte into an anode chamber and a cathode chamber of an electrolysis unit and continually applying direct current to the anode and the cathode of the electrolysis unit to electrolyze, wherein water with the pH value of greater than 7 is produced in the cathode chamber, and a disinfection cleaning solution with the pH value of smaller than 7 is produced in the anode chamber; and mixing the disinfection cleaning solution produced in the anode chamber with the NaCl aqueous solution and filtering with a terminal to obtain the isosmotic disinfection cleaning solution. The preparation method disclosed by the invention is scientific and reasonable, various defects existing in the prior art are overcome, continuous, stable, quick and efficient preparation of the isosmotic disinfection cleaning solution is realized, and the isosmotic disinfection cleaning solution can be applied to cleaning, disinfection and the like of medical instruments and environments such as wounds, wound surfaces, sickrooms, sick beds, clothes, endoscopes, dialysis machines and the like.