text: ISF compounds can be quantified either indirectly from merely diluted ISF samples by using OFM and additional calibration techniques, or directly from undiluted ISF samples which can be collected with additional OFM methods. Quantification of compounds from diluted ISF samples requires additional application of calibration methods, such as Zero Flow Rate, No Net Flux or Ionic Reference. Zero Flow Rate has been used in combination with dOFM by Schaupp et al. to quantify potassium, sodium and glucose in adipose ISF samples. No Net Flux has been applied to quantify several analytes in OFM studies in subcutaneous adipose, muscle and dermal ISF: the absolute lactate concentrations and the absolute glucose concentrations in adipose ISF, the absolute albumin concentration in muscle ISF and the absolute insulin concentration in adipose and muscle ISF have been successfully determined. Dragatin et al. used No Net Flux in combination with dOFM to assess the absolute ISF concentration of a fully human therapeutic antibody. Ionic Reference has been used in combination with OFM to assess the absolute glucose concentration and the absolute lactate concentration in adipose ISF. Dermal OFM has also been used to quantify the concentrations of human insulin and an insulin analogue in the ISF with inulin as exogenous marker.
  Additional OFM methods, such as OFM recirculation and OFM suction can collect undiluted ISF samples from which direct and absolute quantification of compounds is feasible. OFM recirculation to collect undiluted ISF samples recirculates the perfusate in a closed loop until equilibrium concentrations between perfusate and ISF are established. Using albumin as analyte, 20 recirculation cycles have been enough to reach equilibrium ISF concentrations. OFM suction is performed by applying a mild vacuum, which pulls ISF from the tissue into the OFM probe.