BACKGROUND: Annually, infections contribute to approximately 25% of the 2.8 million neonatal deaths worldwide. Over 95% of sepsis-related neonatal deaths occur in low- and middle-income countries. Hand hygiene is an inexpensive and cost-effective method of preventing infection in neonates, making it an affordable and practicable intervention in low- and middle-income country settings. Therefore, hand hygiene practices may hold strong prospects for reducing the occurrence of infection and infection-related neonatal death. OBJECTIVES: To determine the effectiveness of different hand hygiene agents for preventing neonatal infection in both community and health facility settings. SEARCH METHODS: Searches were conducted without date or language limits in December 2022 in the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, Embase and Cumulated Index to Nursing and Allied Health Literature (CINAHL), clinicaltrials.gov and International Clinical Trials Registry Platform (ICTRP) trial registries. The reference lists of retrieved studies or related systematic reviews were screened for studies not identified by the searches.   SELECTION CRITERIA: We included randomized controlled trials (RCTs), cross-over trials, and cluster trials that included pregnant women, mothers, other caregivers, and healthcare workers who received interventions within either the community setting or in health facility settings, and the neonates  in the neonatal care units or community settings. DATA COLLECTION AND ANALYSIS: We used standard methodological procedures expected by Cochrane and the GRADE approach to assess the certainty of evidence. Primary outcomes were incidence of suspected infection (author-defined in study) within the first 28 days of life, bacteriologically confirmed infection within the first 28 days of life, all-cause mortality within the first seven days of life (early neonatal death), and all-cause mortality from the 8th to the 28th day of life (late neonatal death). MAIN RESULTS: Our review included six studies: two RCTs, one cluster-RCT, and three cross-over trials. Three studies involved 3281 neonates; the remaining three did not specify the actual number of neonates included in their study. Three studies involved 279 nurses working in neonatal intensive care units (NICUs). The number of nurses included was not specified by one study. A cluster-RCT included 103 pregnant women of over 34 weeks gestation from 10 villages in a community setting (sources of data: 103 mother-neonate pairs) and another community-based study included 258 married pregnant women at 32 to 34 weeks of gestation (the trial reported adverse events on 258 mothers and 246 neonates). Studies examined the effectiveness of different hand hygiene practices for the incidence of suspected infection (author-defined in study) within the first 28 days of life. Three studies were rated as having low risk for allocation bias,  two studies were rated as unclear risk, and one was rated as having high risk. One study was rated as having a low risk of bias for allocation concealment,  one study was rated as unclear risk, and four werw rated as having high risk. Two studies were rated as having low risk for performance bias and two were rated as having low risk for attrition bias.  One class of agent versus another class of agent: 2% chlorhexidine gluconate (CHG) compared to alcohol hand sanitiser (61% alcohol and emollients) For this comparison, no study assessed the effect of the intervention on the incidence of suspected infection within the first 28 days of life. Two percent chlorhexidine gluconate (CHG) probably reduces the risk of all infection in neonates compared to 61% alcohol hand sanitiser in regard to the incidence of all bacteriologically confirmed infection within the first 28 days of life (RR 0.79, 95% confidence interval (CI) 0.66 to 0.93; 2932 participants, 1 study; moderate-certainty evidence), number needed to treat for an additional beneficial outcome (NNTB): 385. The adverse outcome was reported as mean self-reported skin change and mean observer-reported skin change. There may be little to no difference between the effects of 2% CHG on nurses' skin compared to alcohol hand sanitiser, based on very low-certainty evidence for mean self-reported skin change (mean difference (MD) -0.80, 95% CI -1.59 to 0.01; 119 participants, 1 study) and on mean observer reported skin change (MD -0.19, CI -0.35 to -0.03; 119 participants, 1 study), respectively. We identified no study that reported on all-cause mortality and other outcomes for this comparison. None of the included studies assessed all-cause mortality within the first seven days of life nor the duration of hospital stay. One class of agent versus two or more other classes of agent: CHG compared to plain liquid soap + hand sanitiser We identified no studies that reported on our primary and secondary outcomes for this comparison except for author-defined adverse events. We are very uncertain whether plain soap plus hand sanitiser is better than CHG for nurses' skin based on very low-certainty evidence (MD -1.87, 95% CI -3.74 to -0.00; 16 participants, 1 study; very low-certainty evidence).  One agent versus standard care: alcohol-based handrub (hand sanitiser) versus usual care The evidence is very uncertain whether alcohol-based handrub is better than 'usual care' in the prevention of suspected infections, as reported by mothers (RR 0.98, CI 0.69 to 1.39; 103 participants, 1 study, very low-certainty evidence). We are uncertain whether alcohol-based hand sanitiser is better than 'usual care' in reducing the occurrence of early and late neonatal mortality (RR 0.29, 95% CI 0.01 to 7.00; 103 participants, 1 study; very low-certainty evidence) and (RR 0.29, CI 0.01 to 7.00; 103 participants, 1 study; very low-certainty evidence), respectively. We identified no studies that reported on other outcomes for this comparison. AUTHORS' CONCLUSIONS: We found a paucity of data that would allow us to reach meaningful conclusions pertaining to the superiority of one form of antiseptic hand hygiene agent over another for the prevention of neonatal infection. Also, the sparse available data were of moderate- to very low-certainty. We are uncertain as to the superiority of one hand hygiene agent over another because this review included very few studies with very serious study limitations.