Detecting a gas molecule moving through a small pinhole
The best way to do this is to detect the molecules you want in the presence of others that carry it along. You should not expect to detect every molecule of the type you want but just one of them at a time. 

The most sensitive detection technique is mass spectrometry, more sensitive than fluorescence because if a molecule fluoresces in a direction away from your detector its photon is lost. All ions can in principle be collected in a mass spectrometer.

To get the molecules into the spectrometer mix them with a solvent, (if necessary) and heat this with a pulsed IR laser. This has to be sufficient to vaporise the mixture/substrate  and it has to be positioned close to the input of the spectrometer since this is done outside the spectrometer. The vapour is at atmospheric pressure and mixed with air but because of the pressure differential efficiently enters the spectrometer (it is easy to see this). Here by differential pumping and using skimmers before the entrance to the mass spectrometer part some of the target molecules will be present in the beam produced. The mass spectrometer should probably be of the time of flight/reflectron type to increase throughput and mass resolution. By pulsing the laser it is then possible to gate the detector to observe the molecules at just the right time and so reduce noise. Of course once you have detected the molecule it is destroyed on the detector. 

If you do not want to destroy the molecules then fluorescence detection is the only possibility provided the molecule does fluoresce as not all do with a reasonable yield. A second laser is thus needed after the mass selection and then a photomultiplier is used to count single photons of fluorescence as they arrive (v. common technique now for 50 years or more). I'm not sure what you could do after that but again there is time as photons travel effectively instantaneously compared to the sped the molecule will be travelling at. As I said in a comment not cheap, not trivial.