Nanoprobe arrays for multiple single cell insertion were developed using heterogeneous nanosphere lithography. Using two heterogeneous nanoparticles as sacrificial and masking particles, high aspect ratio Si nanoprobes were fabricated in an array with spacing between the nanoprobes ranging from a few to tens of micrometers. For registered single cell analysis, multiple and precise insertion of nanoprobes into multiple single cells in a parallel fashion was demonstrated using micropipette suction and micromanipulators.