The 3D HSQC-TOCSY experiment is specifically designed to obtain X-edited TOCSY spectra from which ¹H-¹H TOCSY correlations can be clearly assigned even in overcrowded regions (the mechanism involves an heteronuclear transfer via ¹J(XH) followed by a conventional TOCSY step). Similar results can be obtained from an analog 3D HMQC-TOCSY experiment.

Implementation on AVANCE spectrometers is feasible. Improved versions using pulsed field gradients (PFGs) are also available and, therefore, gradient technology is required. The ¹H-¹³C HSQC-TOCSY experiment is usually applied on ¹³C-labeled proteins in D₂O and at natural abundance on concentrated samples. On the other hand, the ¹H-¹⁵N HSQC-TOCSY is usually applied on ¹⁵N-labeled molecules in H₂O.

The original 3D HSQC-TOCSY pulse sequence ( 92JACS1126 ) basically consists of a 2D ¹H-X HSQC pulse sequence followed by an homonuclear 2D TOCSY pulse sequence. Related experiments are:

• The starting HSQC block could be replaced with other heteronuclear schemes as, for instance, the HMQC pulse train (see, for instance, the 3D HMQC-TOCSY experiment).
• The TOCSY block can be replaced by a COSY, ROESY or a NOESY block (see, for instance, the 3D HSQC-NOESY experiment).
• The two building blocks can be reversed (see 3D TOCSY-HSQC experiment)
• A 2D version can be designed by fixing the variable ¹³C evolution period in the HSQC block (2D w1-X-filtered TOCSY experiment) or by fixing the variable ¹H evolution period (see 2D HSQC-TOCSY experiment). In addition, a ¹³C-selective 1D HSQC-TOCSY could be obtained by fixing both variable evolution delays.

Several improved versions have been proposed incorporating the following modifications:

• Improved sensitivity by incorporating the PEP methodology ( 95JMRB170-106 ).
• In the absence of X-decoupling, the 3D HSQC-TOCSY experiment exhibits E.COSY type cross-peaks, allowing the measurement of direct and long-range heteronuclear coupling constants ( 92JACS1126 ).

The 3D HSQC-TOCSY experiment can be recorded in automation mode. Both blocks are usually optimized as in their 2D counterparts.

More details on practical implementation of the 3D HSQC-TOCSY  experiment on AVANCE spectrometers can be found in the corresponding Tutorial 3D HSQC-TOCSY experiment

The HSQC-TOCSY experiment affords a 3D spectrum in which X, ¹H and ¹H chemical shifts are displayed in three independent dimensions. Thus, the F₂(¹H)-F₃(¹H) plane of a ¹H-¹⁵N HSQC-TOCSY spectrum will show a typical 2D F₁(NH)-F₂(¹H) TOCSY map. On the other hand, the F₂(¹H)-F₃(¹H) plane of a ¹H-¹³C HSQC-TOCSY spectrum will show a typical 2D F₁(¹H aliphatic)-F₂(¹H) TOCSY map while the F₁(¹³C)-F₂(¹H) plane will correspond to a conventional 2D ¹H-¹³C HSQC spectrum.