Rapidpoint 400

A venous blood sample was taken to determine plasma levels of sodium, hematocrit, and hemoglobin (Rapidpoint^® 400, Siemens Health Care Diagnostics Inc., Tarrytown, New York) at baseline and directly after exercise cessation. Using changes in plasma hematocrit and hemoglobin concentration the relative plasma volume change (in %) was calculated according to the formula by Dill and Costill (1974). Hypo‐ and hypernatremia were defined as plasma sodium concentrations of ≤135 and ≥145 mmol/L, respectively (Adrogue and Madias 2000; Hew‐Butler et al. 2005). Furthermore, serum creatinine was measured at baseline and directly after completion of the exercise on day 1 and day 3. The estimated glomerular filtration ratio (eGFR) was calculated to assess baseline kidney function using the CKD‐EPI creatinine equation, which contained the serum creatinine concentration as well as subject's age, gender, and ethnicity (Levey et al. 2009). Furthermore, plasma interleukin‐6 (IL‐6) and tumor necrosis factor‐α (TNF‐α) were measured to examine differences in inflammatory response between baseline and postexercise.

Participants were seated for 5 min after which a venous blood sample was taken from the cephalic vein. Blood was collected in a 4 ml Lithium Heparin (LH) gel vacutainer (Becton–Dickinson, New Jersey, America). The vacutainer was centrifuged at 3000G (3755 rpm) for 8 min at 22° and plasma was stored at − 80 °C. Samples were analysed for their iron, ferritin and haptoglobin concentrations in October 2015 (Siemens Dimension Vista 1500, Siemens Healthcare, Erlangen, Germany).
An additional blood sample was collected in a 2 ml LH vacutainer (Becton–Dickinson, New Jersey, USA) and used for direct analyses of plasma haemoglobin and haematocrit concentrations (Rapidpoint 400, Siemens Healthcare Diagnostics Inc., Tarrytown, New York, USA). Relative changes in plasma volume were calculated from blood haematocrit and haemoglobin concentrations using Dill and Costill’s equation (Dill and Costill 1974 (link)). Iron, ferritin, haptoglobin and haemoglobin levels were corrected for plasma volume changes.

Haematology and blood biochemistry were performed at the Clinical Laboratory, Vetsuisse Faculty, University of Zurich (Cases 1–3), using a Sysmex XT-2000iV (Sysmex Corporation, Kobe, Japan) [42 (link)] and a Cobas Integra 800 instrument (Roche Diagnostics AG, Rotkreuz, Switzerland), and at the Clinical Diagnostic Laboratory, Vetsuisse Faculty, University of Bern (Cases 4 and 5), using an Advia 2120 (Siemens Healthcare AG, Zurich, Switzerland) and a Cobas c501 (Roche Diagnostics AG), respectively. Haematocrit and blood biochemistry in Case 5 at first presentation was measured on a Rapidpoint 400 instrument (Siemens Healthcare Diagnostics GmbH, Zurich, Switzerland) and on an IDEXX Vettest Chemistry Analzyer (IDEXX Laboratories, Inc., Westbrook, ME, USA). Modified Wright-Giemsa-stained blood smears (AMES Hema Tek slide stainer, Bayer, Zurich, Switzerland or Hema-Tek 2000, Siemens) were made from fresh EDTA-anticoagulated blood. The Coombs’ test was performed using the microdilution plate method at the Clinical Diagnostic Laboratory, Vetsuisse Faculty, University of Bern using a commercial feline Coombs’ reagent (ImmunO, MP Biomedicals Llc., Solon, OH, USA). The laboratories’ own and published reference intervals [43 (link)] were used for adult cats and published reference intervals were used for kittens [44 (link), 45 (link)].