A new set of genetically encoded calcium indicators (GECOs) has been developed, expanding the color palette for calcium imaging. These indicators include blue, improved green, and red intensimetric indicators, as well as a ratiometric indicator with a 11,000% ratio change. The new indicators enable improved single-color calcium imaging in neurons and transgenic C. elegans. In HeLa cells, calcium was imaged in three subcellular compartments, and calcium and ATP were simultaneously imaged using a cyan FP–yellow FP–based indicator. The development of these indicators was achieved through directed evolution, which involved screening large libraries of genetic variants to identify those with improved properties. The most optimized single FP calcium indicator, GCaMP3, was used as a starting point for the evolution process. The resulting G-GECO1, G-GECO1.1, and G-GECO1.2 indicators show a significant increase in fluorescence compared to GCaMP3. Additionally, blue (B-GECO), red (R-GECO), and ratiometric (GEM-GECO) indicators were developed, each with unique properties. The GEM-GECO1 indicator exhibits a large ratiometric change and is useful for imaging calcium in whole animals. The R-GECO1 indicator shows improved sensitivity for imaging neuronal activity in C. elegans. The new indicators also enable multicolor calcium imaging in single cells, allowing for the visualization of correlated changes in calcium in different organelles. The development of these indicators opens up new possibilities for calcium imaging experiments that were previously impractical. The indicators enable imaging of multiple calcium indicators in single cells, improved sensitivity for imaging neuronal activity in C. elegans, and multiparameter imaging with CFP-YFP FRET-based biosensors. Additionally, GEM-GECO1 and R-GECO1 should facilitate imaging of neuronal activity after optogenetic channel activation.A new set of genetically encoded calcium indicators (GECOs) has been developed, expanding the color palette for calcium imaging. These indicators include blue, improved green, and red intensimetric indicators, as well as a ratiometric indicator with a 11,000% ratio change. The new indicators enable improved single-color calcium imaging in neurons and transgenic C. elegans. In HeLa cells, calcium was imaged in three subcellular compartments, and calcium and ATP were simultaneously imaged using a cyan FP–yellow FP–based indicator. The development of these indicators was achieved through directed evolution, which involved screening large libraries of genetic variants to identify those with improved properties. The most optimized single FP calcium indicator, GCaMP3, was used as a starting point for the evolution process. The resulting G-GECO1, G-GECO1.1, and G-GECO1.2 indicators show a significant increase in fluorescence compared to GCaMP3. Additionally, blue (B-GECO), red (R-GECO), and ratiometric (GEM-GECO) indicators were developed, each with unique properties. The GEM-GECO1 indicator exhibits a large ratiometric change and is useful for imaging calcium in whole animals. The R-GECO1 indicator shows improved sensitivity for imaging neuronal activity in C. elegans. The new indicators also enable multicolor calcium imaging in single cells, allowing for the visualization of correlated changes in calcium in different organelles. The development of these indicators opens up new possibilities for calcium imaging experiments that were previously impractical. The indicators enable imaging of multiple calcium indicators in single cells, improved sensitivity for imaging neuronal activity in C. elegans, and multiparameter imaging with CFP-YFP FRET-based biosensors. Additionally, GEM-GECO1 and R-GECO1 should facilitate imaging of neuronal activity after optogenetic channel activation.