Figure 1. Example data for optimizing the choice of four objects in C57/129J mice. Starting from two previously tested objects (dome and steps), we proceeded to add two new objects with the goal of finding a non-biased four-object combination. When finding innate preference for one or more objects (A), possible strategies are to (1) modify features that drive bias (B; for example, by removing the flat top in the flat hourglass design); or (2) replace preferred objects until finding an ideal object combination (C; for example, we introduced two novel designs, star and icosahedron, but found innate preference for the star object; this was then followed by D, when we replaced the star with the steps object to finally see an absence of innate preference). Multiple rounds of tests may be necessary, but in our experience, innate preferences are quite robust across animal cohorts in future experiments. Bars represent mean exploration time ± SEM. HG, hourglass; *p < 0.05. Methodology: adult C57/129J (a cross between C57BLK/6J and 129S1/SvImJ strains) mice were handled and habituated (5 min of handling followed by placement into the behavioral chamber for 4 min) to the square open-field behavioral chamber (30 × 30 × 30 cm) twice a day for four consecutive days. On the following day, mice were placed back into the chamber to interact for 5 min with four objects located 3 cm away from each wall at each corner of the chamber. Exploration time (seconds) per object was measured using ANY-maze software. Object location and start position within the chamber were counterbalanced and randomized across subjects. All animal procedures were approved by the local Animal Care Committee. Statistics: (A) one-way RM ANOVA F(3,15) = 7.53, p = 0.011; Tukey’s post hoc test shows a significant difference between steps and flat HG p = 0.04; (B) one-way RM ANOVA F(3,57) = 9.10, p = 0.0003; Tukey’s post hoc test shows a significant difference between pyramid versus steps p = 0.042, pyramid versus angled HG p = 0.0004, dome versus steps p = 0.031, dome versus angled HG p = 0.0003; (C) one-way RM ANOVA F(3,36) = 5.59, p = 0.003; Tukey’s post hoc test shows a significant difference between dome versus star p = 0.002, star versus icosahedron p = 0.036; (D) one-way RM ANOVA F(3,27) = 1.51, p = 0.25.