Amblyopia is defined as impaired visual acuity in one or both eyes without demonstrable abnormality of the visual pathway, and is not immediately resolved by wearing glasses. In performing this systematic review, we aimed to synthesize the best available evidence regarding the effectiveness and safety of conventional occlusion therapy compared to atropine penalization in treating amblyopia. We searched the Cochrane Central Register of Controlled Trials (CENTRAL) (which contains the Cochrane Eyes and Vision Trials Register) (2018, Issue 8); Ovid MEDLINE; Ovid Embase; LILACS BIREME; ClinicalTrials.gov; ISRCTN; and the WHO ICTRP on 7 September 2018. We included randomized/quasi-randomized controlled trials comparing conventional occlusion to atropine penalization for amblyopia. Two review authors independently screened abstracts and full-text articles, abstracted data, and assessed risk of bias. We included seven trials (five randomized controlled trials and two quasi-randomized controlled trials) conducted in six countries (China, India, Iran, Ireland, Spain, and the United States) with a total of 1177 amblyopic eyes. Three of these seven trials were from the original 2009 version of the review. We assessed two trials as having a low risk of bias across all domains, and the remaining five trials as having unclear or high risk of bias for some domains.As different occlusion modalities, atropine penalization regimens, and populations were used across the included trials, we did not conduct any meta-analysis due to clinical and statistical heterogeneity. Evidence from six trials (two at low risk of bias) suggests that atropine penalization is as effective as conventional occlusion in improving visual acuity. Similar improvement in visual acuity was reported at all time points at which it was assessed, ranging from five weeks (improvement of 1 line) to 10 years (improvement of greater than 3 lines). At six months, although most participants (363/522) come from a trial rated as at low risk of bias with a precise estimate (mean difference (MD) 0.03, 95% confidence interval (CI) 0.00 to 0.06), two other trials rated as at high risk of bias produced inconsistent estimates and wide confidence intervals (MD -0.02, 95% CI -0.11 to 0.07 and MD -0.14, 95% CI -0.23 to -0.05; moderate-certainty evidence). At 24 months, additional improvement was found in both groups, but there continued to be no meaningful difference between those receiving occlusion and those receiving atropine therapies (moderate-certainty evidence).We did not find any difference in ocular alignment, stereo acuity, or sound eye visual acuity between occlusion and atropine penalization groups (moderate-certainty evidence). Both treatments were well tolerated. Atropine was associated with better adherence (moderate-certainty evidence) and quality of life (moderate-certainty evidence), but also a higher reported risk of adverse events in terms of mild reduction in the visual acuity of the sound eye not requiring treatment and light sensitivity (high-certainty evidence). Skin, lid, or conjunctival irritation were more common among participants receiving patching than those receiving atropine (high-certainty evidence). Atropine penalization costs less than conventional occlusion. Both conventional occlusion and atropine penalization produce visual acuity improvement in the amblyopic eye. Atropine penalization appears to be as effective as conventional occlusion, although the magnitude of improvement differed among the trials we analyzed.