Implementation and evaluation of a detector for forward propagated second harmonic signals

Second harmonic generation (SHG) is emerging as an alternative non-linear imaging method. The fact that most commercial multi-photon microscopes can be easily adapted to image SHG makes it appealing to explore the kind of sample information given by SHG. Here we describe an SHG detection implementation designed to optimize the collection of forward propagating light. A Hamamatsu H957-08 PMT is inserted at the back-focal plane of the condenser on an inverted Nikon TE300, and controlled by the existing electronics of a BioRad 1024MP system. Evaluation of the performance was done on common SHG generating preparations, KH2PO4 crystals and collagen. We concluded that positioning a detector at the back focal plane of the condenser provides a highly efficient detection system for second harmonic signals, with many advantages over a detector sited at the lamp housing. (C) 2004 Elsevier Ltd. All rights reserved.