Various Gram staining automated systems are available to accelerate and standardize the staining process but a systematic comparison of different systems is largely lacking. The objective of this study was to evaluate two devices in comparison to manual Gram staining. Clinical samples (n=500, University Hospital Münster, Germany, May-June 2020) were simultaneously Gram stained manually and with two automated Gram strainers (PREVI® Color Gram, bioMérieux and ColorAX2, Axonlab). The quality was assessed based on four criteria: (i) homogeneous staining of bacteria/fungi, (ii) uniform staining of the background, (iii) absence of staining artifacts and (iv) congruency between culture and microscopy. Each criterion was rated with “0” (absence) or “1” (presence) point to calculate a quality score (0–4 points). The costs for each staining procedure were calculated based on consumables and hands-on time (applying the average wage of a laboratory technician in the public service for Germany and the US). The mean (±SD) quality scores were comparable for manual staining (3.06±0.91) and PREVI® Color Gram (3.04±0.90, p=0.6) while significantly lower scores were achieved by ColorAX2 (2.57±1.09, p<0.0001). The total cost per Gram staining was 1.13 €/1.34 $ (PREVI® Color Gram), 0.80 €/0.83 $ (manual) and 0.60 €/0.71 $ (ColorAX2), respectively. The quality and costs per slide vary significantly between instruments of different manufacturers.