The present invention provides human protein arrays that include at least 1000 human proteins. In another embodiment, the present invention provides a method for identifying a substrate of an enzyme, comprising contacting the enzyme with a positionally addressable array comprising at least 100 proteins immobilized on functionalized glass surface, and identifying a protein on the positionally addressable array that is bound and/or modified by the enzyme, wherein a binding or modifying of the protein by the enzyme indicates that the protein is a substrate for the enzyme. In additional embodiments, provided herein are methods for making an array of at least 1000 human proteins under non-denaturing conditions, including human proteins that are difficult to express and/or difficult to isolate in a non-denatured state.