Over the last decade microfluidics has created a versatile platform that has significantly advanced the ways in which micro-scale organisms and objects are controlled, processed and investigated, by improving the cost, compactness and throughput aspects of analysis. Microfluidics has also expanded into optics to create reconfigurable and flexible optical devices such as reconfigurable lenses, lasers, waveguides, switches, and on-chip microscopes. Here we present a new opto-fluidic microscopy modality, i.e., Holographic Opto-fluidic Microscopy (HOM), based on lensless holographic imaging. This imaging modality complements the miniaturization provided by microfluidics and would allow the integration of microscopy into existing on-chip microfluidic devices with various functionalities. Our imaging modality utilizes partially coherent in-line holography and pixel super-resolution to create high-resolution amplitude and phase images of the objects flowing within micro-fluidic channels, which we demonstrate by imaging C. elegans, Giardia lamblia, and Mulberry pollen. HOM does not involve complicated fabrication processes or precise alignment, nor does it require a highly uniform flow of objects within microfluidic channels.