The scale of entrepreneurial ecosystems (EE) assesses the perceptions about entrepreneurial ecosystem domains, finances, capital finances, support, support professions, policies, markets, human resources, and culture. The scales are always error-prone—these scales must possess properties that enable them it to provide maximum information and validity reliability. Convenient sampling data from (n = 474) founders, co-founders, and entrepreneurs were collected. The IRT-GRM model is used to validate and test the instrument-based on polytomous scales. IRT yields discriminating power—the level of difficulty of the items of the scale. The scale consists of 48 items. The item Pol5 (4.13) was found to have the highest discriminating value (4.13), the item mar5 had the lowest discriminating value (1.57), and all items had discriminating values greater than the threshold value of 0.60. The EE Scale showed good reliability based on McDonald’s omega and Cronbach’s alpha (0.80 and 0.88). The parallel and factor analysis showed good agreement of the one-dimesnionality of the scale. The model goodness of fit statistics based on the comparative fit index (CFI) and the Tucker–Lewis index, (TLI) and the standardized root mean square residual (SRMR) showed a satisfactory level of fit; however, the root mean square error of approximation (RMSE) showed a poor fit. The item characteristic curves showed that the all item responses were properly ordered. The items of the scale showed a satisfactory level of discrimination power and level of difficulty, and it was found to have three levels of agreement about entrepreneurial ecosystem scale. It is concluded that the EE scale possesses good psychometric properties and that it is reliable and valid instrument to measure the entrepreneurial ecosystem of the given region.