SPDM Localisation Microscopy

SPDMphymod Blinking for standard dyes

 

 Spectral Precision Distance Microscopy, the first described localization microscopy technology 1997 is a light optical process of fluorescence microscopy, which allows position, distance and angle measurements on "optically isolated" particles (e.g. molecules) well below the theoretical limit of resolution for light microscopy.

 

"Optically isolated" means that at a given point in time, only a single particle/molecule within a region of a size determined by conventional optical resolution (typically approx. 200-250 nm diameter is being registered.  This is possible when molecules within such a region all carry different spectral markers (e.g. different colors or other usable differences in the light emission of different particles).


The structural resolution achievable using SPDM can be expressed in terms of the smallest measurable distance between two in their spatial position determined punctiform particle of different spectral characteristics ("topological resolution").  Modeling has shown that under suitable conditions regarding the precision of localization, particle density etc., the "topological resolution" corresponds to a "space frequency," which in terms of the classical definition is equivalent to a much-improved optical resolution.

 

SPDM is a localization microscopy that achieves an effective optical resolution several times better than the conventional optical resolution, represented by the half-width of the main maximum of the effective point image function.  By applying suitable laser optical precision processes, position and distances can be measured with nanometer accuracy between targets with different spectral signatures.

 

References

  • US patent 6,424,421, filed 23rd December 1996: Christoph Cremer, Michael Hausmann, Joachim Bradl, Bernd Rinke ''Method and devices for measuring distances between object structures''
  • J. Bradl, B. Rinke, A. Esa, P. Edelmann, H. Krieger, B. Schneider, M. Hausmann, C. Cremer (1996): ''Comparative study of three-dimensional localization accuracy in conventional, confocal laser scanning and axialtomographic fluorescence light microscopy''. Proc. SPIE 2926: 201-206
  • R. Heintzmann, H. Münch, C. Cremer (1997) ''High-precision measurements in epifluorescent microscopy - simulation and experiment''. Cell Vision 4: 252-253
  • Lemmer P, Gunkel M, Baddeley D, Kaufmann R, Urich A, Weiland Y, Reymann J,  Müller P, Hausmann M, Cremer C (2007): ''SPDM – Light Microscopy with Single Molecule Resolution at the Nanoscale''. In: Applied Physics B  2008; 93: 1–12