We detect by optical microscopy Au and fluorescent nanopartides (NPs) during their motion in water based medium, using an array of dielectric microspheres that are patterned in a microwell array template. The Microspheres act as lenses focusing the light originating from a microscope objective into so-called photonic nanojets that expose the medium Within a microfluidic channel When a NP is randomly transported through a nanojet, its backscattered light (for a bare Au NP) or its fluorescent emission is instantaneously detected by video microscopy. Au NPs down to 50 nit in size, as well as fluorescent NPs down to 20 rim in size, are observed by using a low magnification/low numerical aperture microscope objective in bright field or fluorescence mode, respectively. Compared to the NPs present outside of the photonic nanojets, the light scattering or fluorescence intensity of the NPs in the nanojets is typically enhanced by tip to a factor of The experimental intensity is found to be proportional to the area Occupied by the NP in the nanojet. The technique is also used for immunodectection of biomolecules immobilized on At NPs in buffer and; in future, it may develop into a versatile tool to detect nanometric objects of environmental or biological importance, such as NPs, viruses, or other biological agents.