The National Maternal and Child Health Center in Phnom Penh, Cambodia commenced PMTCT services as the first site of the National PMTCT programme of Cambodia in November 2001. However, the acceptance of voluntary confidential counselling and testing (VCCT) in the PMTCT services was not as high as expected. The aim of this study was to evaluate influence of partner participation in the mother class to the PMTCT services. During the first visit to antenatal care, all women were invited to the class, where information on PMTCT was provided. From July 2002, when partner participation started, to May 2005, the acceptance rate to the pre-test counselling of those who attended the class alone was 18.7% (3,234/17,340), while that of the attendees with their partner was 85.1% (2,908/3,417) (p <0.001). All of the couples accepted couple counselling and disclosure of their results to their partners. In conclusion, a strong association was observed between acceptance and partner involvement. However, we should consider also other strategies for future programme improvement.