We demonstrate the fabrication of good quality surface alignment layers on glass by Direct Laser Writing method using a 2-photon polymerisation technique. We use commercially available photosensitive resins to print alignment layers by scanning the focal point of a femtosecond laser near the resin-glass interface. This results in down to ~ 100 nm thin alignment layers that provide good planar anchoring of 5CB and MLC13300, with the easy axis of alignment along the scanning direction. The azimuthal anchoring strength is ~ 5 × 10-6 J/m2 and is an order of magnitude weaker compared to commercial rubbed polyimide alignment layer. The threshold voltage for Fréedericksz transition in a 90° twisted nematic cell is slightly increased compared to conventional rubbed polyimide for printed alignment layers. The turn-on switching time is longer for printed layers compared to polyimide alignment layers, whereas the turn-off time is shorter for printed alignment layers. The advantage of this new method is in its flexibility, as we demonstrate printing of complex surface alignment patterns with alignment layer thickness below 100 nm.