Middlesex Township lies entirely within the "Last Purchase of 1784" being the next to last land bought from the Indians as negotiated by the second Treaty of Ft. Stanwix.Middlesex Township was one of the four original townships recognized as early as 1794 still located in Allegheny County, and one of the original thirteen townships of Butler County in 1804.By Act of Legislation on March 12, 1800 the County of Butler was officially born. This Act provided for the attachment of Butler County to Allegheny County for administrative purposes. The Act also provided that the county seat be located no further than four miles from the center of the county.

Thus, in March 1803, three men were authorized to survey 300 acres on the north side of the Connoquenessing Creek and lay out lots for sale, setting aside a piece of land not exceeding five acres for county buildings. From 1800 to the end of 1803, a total of $5,5283.90 in taxes was collected from the original four townships of Connoquenessing, Middlesex, Slippery Rock, and Buffalo.Middlesex Township leaders submitted a petition to the Court on June 18, 1853 leading to the ultimate establishment of today's township boundaries. The state finalized the matter by an Act of Legislature March 29, 1854 and today's Middlesex Township was born. The Courts approved the present division of thirty-three Townships, each one being approximately five miles square.

It has been theorized that the name "Middlesex" was derived from the Township's position in between or in the middle of other municipalities. It lies along the Allegheny County line, bordered by Adams Township to the west, Forward Township to the northwest, Penn Township to the north, Richland Township, Allegheny County to the south, and Clinton Township to the east.The southeastern corner of the Township has always seemed to be the Townships' "cradle of civilization." Many early industries included a gristmill, sawmill, tannery, distillery, and a coffin shop in 1822.Most of the Township's newcomers settle in the southern parts of the Township, while the north is still primarily farmland.