Reference tissue

Reference tissue is a region where

Reference tissue can be used as a substitute of arterial plasma curve as input function in quantitative analysis of PET studies, applying compartmental models or multiple-time graphical analyses, or in semi-quantitative analysis applying tissue-to-reference tissue ratio. Ex vivo studies are needed to validate that reference tissue has no specific binding of the ligand.

In optimal situation the ratio of reference tissue and plasma input curves reaches a steady level, and may even be used to estimate the fractions of parent tracer and labelled metabolites in the plasma (Wong et al., 1986).

Reference tissue vs arterial plasma as input function

Reference tissue Arterial plasma
  • noninvasive method
  • can provide binding potential in receptor studies
  • optimal reference tissue is available only for a few brain receptor tracers
  • radioactivity spillover from adjacent tissues or due to scatter
  • specific analysis models required
  • represents the true input function (gold standard)
  • can be corrected for labelled metabolites
  • invasive
  • not accessible from all subjects or small animals
  • labour-intensive and error-prone metabolite analysis

If reference tissue has specific binding

If the reference region has specific binding, the binding potential will be underestimated (Gunn et al. 1997):

Specific binding in reference region will also lead to bias in Scatchard analysis (Litton et al. 1994). However, correction methods have been developed (Turkheimer et al., 2012).

Specific binding in reference tissue may lead to severe underestimation of receptor occupancy.

If reference tissue contains labelled metabolite

If label-carrying metabolite passed the blood-brain barrier, then reference tissue input model may lead to severe bias in results, and plasma input models may be preferable (Zoghbi et al., 2006). Specific model needs to be developed, where the concentrations of the metabolite in plasma is used as second model input (Matsubara et al., 2014).

For certain tracers it may be possible that the concentration of labelled metabolite at late time points is minimal and can be ignored (Kim et al., 1999).

Extracerebral reference region

When no suitable cerebral reference region is available, it may be possible to quantify brain receptors using muscle as reference region (Le Foll et al. 2007). However, there are several caveats in using this approach:

  1. Radioactive label carrying metabolites that can not penetrate the blood brain barrier may well enter extracellular tissues and prevent their use as reference region
  2. Blood flow is much lower in muscle than in the brain, therefore becoming the limiting factor for K1 in the reference tissue
  3. Furthermore, blood flow in muscle is highly variable in awake state
  4. Non-displaceable distribution volume is probably different in the brain and in extracerebral reference region; therefore it has to be measured, and only if proven to be constant the binding potentials can be corrected for it (Le Foll et al. 2007)
  5. Additional requirements may be set by the model, for example, SRTM requires that reference region kinetics can be reasonably well described by 1-tissue compartment model
Tumour In oncology, the concentration in tumour is frequently divided by concentration in normal tissue, which often is muscle (T/N or T/M). This mimics visual analysis, but gives a numerical value to the target-to-background difference.
Problem is that radioligand uptake in muscle may multiply if patient is nervous or position is hard to keep.

Tissue-to-plasma ratio may be a better alternative in these cases, if at least one or two (arterialized) venous blood samples can be taken during the late-scan.

“Positive” reference tissue

If radioligand uptake is very high in a certain brain region, it may take up all ligand that is available in the plasma. In that case the tissue curve is proportional to the AUC (integral) of arterial plasma curve. “Positive” reference tissue can be found and used with few radioligands, including certain AChE tracers.

See also:


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Updated at: 2018-02-14
Written by: Vesa Oikonen