In vitro carrier tests, suspension tests, time-kill curves, and determinations of minimum inhibitory concentrations to evaluate the microbicidal activities of hand antiseptics provide only a preliminary indication of the antimicrobial spectrum and speed of action of a given formulation.

Ex vivo testing on human or animal skin at human skin temperature and at contact times reflecting field conditions may give a better indication of a formulation’s ability to eliminate hand-transmitted pathogens.

European Norms exist for assessing the antimicrobial efficacy of hand disinfectants.

A suspension test for the demonstration of bactericidal activity (EN 12054) is obligatory for hand disinfectants in all fields of application; a test to prove activity against yeasts applies only to hygienic hand rub. Claims for virucidal activities should be substantiated  by EN 1476. In vivo tests exist for post-contamination treatments, hygienic hand wash and hygienic hand rub (EN 1499 and EN 1500 respectively).

All in vivo tests use reference hand treatments (with unmedicated soap or 2-propanol 60% (vol.) or 1-propanol 60% (vol.), respectively), the results of which are compared with those achieved with the product under test and with the same volunteers.

An antiseptic soap needs to be significantly more efficacious than unmedicated soap, a product for hygienic hand rub must not be inferior to the reference treatment with 2-propanol.

In the USA, antiseptic preparations intended for use as surgical hand preparation, based on the Food and Drug Administration (FDA) Tentative Final Monograph, are evaluated for their ability to reduce the number of bacteria released from hands immediately after scrubbing, after wearing surgical gloves for 6 h (persistent activity), and after multiple applications over five days (cumulative activity). The most important criteria are immediate and persistent activity. US guidelines recommend that agents for surgical hand preparation should significantly reduce micro-organisms on intact skin, have broad-spectrum activity, and be fast-acting and persistent.

In Europe, all products must be at least as efficacious as a reference surgical rub with n-propanol as outlined in the European Norm EN 12791. By contrast to the USA, only the immediate effect after the hand hygiene procedure and the level of regrowth after 3 h under the gloved hands are measured. The cumulative effect over five days is not a requirement of EN 12791.

Among surgical hand antisepsis using medicated soap the most commonly used products for surgical hand antisepsis are chlorhexidine or povidone-iodine-containing soaps. The most active agents (in order of decreasing activity) are chlorhexidine gluconate, iodophors, triclosan, and plain soap.

Application of chlorhexidine or povidone-iodine results in similar initial reductions of bacterial counts (70-80%) that increase to 99% after repeated applications. Rapid regrowth occurs after the application of  povidone-iodine, but not after the use of chlorhexidine.

Hexachlorophene and triclosan detergents show a lower immediate reduction, but a good residual effect. These agents are no longer commonly used in operating theatres because other products such as chlorhexidine or povidone-iodine provide similar efficacy at lower levels of toxicity, faster mode of action, or broader spectrum of activity.

Povidone-iodine remains one of the most widely used products for surgical hand antisepsis, despite both in-vitro and in-vivo studies demonstrating that it is less efficacious than chlorhexidine, induces more allergic reactions, and does not show similar residual effects.

Awareness has recently arisen that the current tests do not really resonate with the real world regarding the amount of rubs that are used and the speed of reduction of organisms required, and methods for relating the effectiveness of antibacterial handwashing products to infection reduction and for conducting future studies designed to demonstrate the efficacy of antibacterial handwash formulations are needed.