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Stability of chemical and immunochemical analytes in uncentrifuged plasma

samples

Article  in  Annals of Clinical Biochemistry · January 2009

DOI: 10.1258/acb.2008.008212 · Source: PubMed

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Turku University Hospital Helsinki University Central Hospital
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Short Report

Stability of chemical and immunochemical analytes

in uncentrifuged plasma samples

Aila Leino1,2 and M K Koivula1

1
TYKSLAB, the Hospital District of Southwest Finland; 2Department of Clinical Chemistry, University Hospital, Kiinamyllynkatu 4-8,
FIN-20520, Turku, Finland
Corresponding author: Aila Leino. Email aila.leino@tyks.fi

Abstract
Background: The stability of analyte concentrations in plasma after prolonged contact with blood cells in uncentrifuged
lithium-heparin gel tubes was studied.
Methods: To investigate the stability of concentrations of 26 chemistry and 15 immunochemistry analytes, the simultaneously
drawn samples (n ¼ 50) were measured after 6 h storage at þ88C and þ228C in whole blood and after immediate separation of
plasma. The analyte concentrations were measured with a Roche Modular PPEE analyser using reagents from Roche Diagnostics.
Results: After prolonged contact with cells a clinically significant change was only observed for potassium where the mean value
increased from 4.0 mmol/L to 4.8 mmol/L (P , 0.001) when stored at þ88C.
Conclusion: Immediate separation of plasma from cells is recommended. However, when prolonged contact of plasma with cell
is unavoidable, samples can be kept uncentrifuged for up to 6 h at þ88C or at þ228C. The stability of potassium, however,
is temperature-dependent and cannot be measured from refrigerated blood samples.

Ann Clin Biochem 2009; 46: 159– 161. DOI: 10.1258/acb.2008.008212

Introduction tubes (Terumo Europe, Belgium). Cell-free plasma was

Plasma-based testing has increased in many laboratories to
reduce turnaround time by shortening the time interval
between blood collection and testing. However, a common
problem in the daily routine facing clinical laboratories is
the uncertainty of the integrity of uncentrifuged specimens
for chemical and immunochemical analyses. It is very well
known that, ideally, the plasma should be separated from
cells as quickly as possible to prevent the ongoing metab-
olism of cellular constituents.1 However, the allowance of
patient specimens to prolonged contact with blood cells
would offer more flexibility to unload the early morning
rush hours in many laboratories and loosen the requirements
of transportation of specimens to testing laboratories.
Publications on analyte stability in uncentrifuged heparin
plasma specimens are few and concern mainly the stability
of the concentrations of chemical analytes.1 – 4
In this study we determined the stability of concentrations
of 26 chemical and 15 immunochemical analytes after 6 h of
storage in whole blood. In addition, the influence of storage
temperature and transportation (courier or bus) of the tubes
on analytical stability was examined.
Materials and methods
Venous blood was simultaneously collected from 50 fasting
volunteers into three 3.5 mL VenoSafeTM lithium heparin gel
obtained after 6 h storage at þ228C and þ88C in whole
blood and after immediate (within 0.5 h) separation of
plasma. All plasma samples were centrifuged at 2500 g for
10 min at þ188C according to manufacturer’s instructions
(Terumo Europe, Belgium).
All analytes were measured from a single tube with one
Roche Modular PPEE analyser, with commercial reagents pro-
vided by Roche Diagnostica (Roche Diagnostics, Germany).
Thus, components such as glucose, demanding a special
blood sampling tube for a longer stability were not considered.
Statistically significant changes were determined for each
analyte by one-way analysis of variance (ANOVA) or
Kruskal-Wallis test. Statistical analyses were performed
using SPSS 16.0 (SPSS Inc, Chicago, IL, USA).
To determine the changes in analyte concentrations of
plasma with and without prolonged contact with cells, the
mean from 50 volunteers for each respective analyte was
obtained. Clinically significant changes were determined
by the significant change limit5 (SCL) approach defined as:
SCL ¼ Initial value + 2.8 usual SD.
It is based on the assumption that the usual SD (USD) is
representative of the inherent day-to-day variability of the
method. In this study, the calculated mean for each
analyte at 0.5 h represented the initial value. USD was
obtained by averaging the SD of the quality-control data

Annals of Clinical Biochemistry 2009; 46: 159– 161

160 Annals of Clinical Biochemistry Volume 46 March 2009
................................................................................................................................................

(target mean most closely matched the 0.5 h mean) observed
for the last 12 months for each respective analyte.5 SCL was
calculated by establishing the range (+2.8 USD) from mean
at 0.5 h.
Results and discussion
Table 1 shows the statistics for plasma analyte concentrations
obtained in samples separated immediately after collection,
along with those of 6 h storage as whole blood. The results
indicate that the concentrations of 25 chemical and 15
immunochemical analytes were quite stable when samples
were stored uncentrifuged for 6 h either at þ228C or þ88C.
Only for potassium did the observed mean exceeded the
SCL, increasing from 4.0 mmol/L to 4.8 mmol/L (P ,
0.001) when stored at þ88C. This is known to be due to the
Naþ, Kþ-ATPase activity brought about by cold.6 Our
results are in accordance with the recently published study
of Tanner et al.7 although their findings are based on serum.
In order to study the effect of transportation (courier or
bus) on the stability of analyte concentrations, the con-
ditions were simulated by strong-handed manipulation of
the tubes. Neither the continuous gentle swinging in a
mixer nor substantial upside down turning of the

Table 1 Stability of plasma analytes after 6 h storage in whole blood

Storage time and temperature
Analyte (units) Range 0.5 h at 12288 C, mean† 6 h at 12288 C, mean† 6 h at 1888 C, mean† USD value SCL range‡
P –ALP (U/L) 14– 1202 114 113 114 4 100– 124
P –ALB (g/L) 16– 45 33 33 33 0.77 32– 36
P –ALT (U/L) 8– 393 48 50 49 2 41– 53
P –AMYL (U/L) 5– 172 53 53 53 3 44– 62
P –BIL (mmol/L) 4– 350 27 27 27 1 23– 29
fP– Ca (mmol/L) 1.84 – 2.47 2.22 2.23 2.20 0.040 2.10 – 2.34
P –CK (U/L) 5– 1042 118 120 117 5 102– 132
P –Cl (mmol/L) 90– 110 104 103 103 2 98– 110
P –CRP (mg/L) 0– 305 43 44 43 2 35– 47
P –Fe (mmol/L) 2– 35 10.8 11.6 11.1 0.5 9.5 – 12.5
P –GGT (U/L) 6– 1075 100 100 100 3 87– 105
P –HAPTO (g/L) 0– 4.6 2.0 2.0 2.0 0.08 1.7 – 2.1
P –K (mmol/L) 3.0– 5.6 4.0 4.0 4.8§ 0.08 3.7 – 4.1
P –CHOL (mmol/L) 1.9– 9.7 4.1 4.1 4.0 0.15 3.7 – 4.6
P –CHOL – HDL (mmol/L) 0.21 – 3.10 1.14 1.14 1.10 0.040 1.02 – 1.26
fP– CREA (mmol/L) 46– 194 83 84 84 4 71– 95
fP– LDH (U/L) 101– 831 264 276 282 9 237– 291
P –Mg (mmol/L) 0.65 – 1.12 0.95 1.01§ 0.97 0.050 0.80 – 1.10
P –Na (mmol/L) 127– 146 139 140 138 1.5 135– 144
fP– PHOS (mmol/L) 0.67 – 1.59 1.09 0.96§ 1.12 0.05 0.95 – 1.25
P –RF (IU/mL) 0– 121 4 4 4 1.0 4– 10
P –Transferrin (g/L) 0.9– 3.2 2.1 2.1 2.1 0.12 1.9 – 2.6
P –TfR (mg/L) 0.7– 5.7 2.0 2.0 2.0 0.15 1.6 – 2.5
fP– TG (mmol/L) 0.53 – 5.21 1.47 1.47 1.48 0.1 1.17 – 1.77
P –UA (mmol/L) 117– 861 359 355 361 7 338– 380
P –BUN (mmol/L) 1.7– 19.3 6.3 6.2 6.2 0.25 5.5 – 7.0
P –B12 ( pmol/L) 149– 1476 514 490 510 30 416– 596
P –CA12 – 5 (kU/L) 3– 818 57 57 57 2.7 49– 65
P –CA15 – 3 (kU/L) 6– 44 16 16 16 1.3 12– 20
P –CA19 – 9 (kU/L) 1– 350 22 22 22 1.2 18– 26
P –CEA (mg/L) 0.2– 5.7 2.1 2.2 2.1 0.30 1.2 – 3.0
P –CK –MBm (mg/L) 1.1– 90.7 5.2 5.2 5.2 0.23 4.4 – 5.8
fP– C– PEPT (nmol/L) 0.3– 4.1 1.2 1.2 1.2 0.08 0.1 – 1.4
fP– INSU (mU/L) 1– 478 28 26 27 1.3 23– 31
P –CORSOL (nmol/L) 181– 1750 503 504 512 15 444– 534
fP– PTH (ng/L) 24– 187 61 57 60 3.8 50– 72
P –PSA (mg/L) 0– 21.7 2.7 2.7 2.7 0.18 2.3 – 3.3
P –FPSA (mg/L) 0– 2.0 0.40 0.38 0.39 0.02 0.34 – 0.46
P –TnT (mg/L) 0.01 – 4.22 0.36 0.36 0.36 0.080 0.10 – 0.58
P –TSH (mU/L) 0.50 – 13.03 2.15 2.15 2.15 0.270 1.54 – 3.16
P –FT4 ( pmol/L) 10.1 – 24.3 16.0 16.5 16.4 1.24 12.2 – 19.6

SCL, significant change limit; USD, usual standard deviation; ALP, alkaline phosphetase; ALB, albumin; ALp, alanine transaminase; AMYL, amylase; BIL, billrubin;
Ca, calcium; CK, creatinine kinase; CI, chloride; CRP, c-reactive protein; Fe, iron; GGT, gamma glutamyl transferase; HAPTO, haptoglobin; K, potassium; CHOL,
cholesterol; CREA, creatinine; LDH, lactate hydrogenase; Mg. magnesium; Na, sodium; PHOS, inorganic phosphate; RF, rheumatoid factor; TRANSF, transferrin;
TfR, transferrin receptor; TG, triglycerides; UA, uric acid; BUN, urea; B12, vitamin B12; CEA, carcino embryonal antigen; C-PEPT, C-peptide; INSU, insulin;
CORSOL, cortisol; PTH, parathormone; PSA, prostate specific antigen; FPSA, free PSA; TnT, troponin T; TSH, thyroid-stimulating hormone; FT4, free T4

Range of results from 0.5 h concentration at þ228C (n ¼ 50)
†
Means of results (n ¼ 50)
‡
Mean at 0.5 h+2.8 USD
§
Statistically significant difference from 0.5 h concentration (one-way analysis of variance or Kruskal-Wallis test, P , 0.05)

SCLs have been exceeded
 Leino and Koivula. Chemical and immunochemical analytes in uncentrifuged plasma samples
................................................................................................................................................
uncentrifuged tubes during the prolonged storage had an
additional effect on measured concentrations of analytes.
Our results indicate that in the case when lithium-heparin
plasma specimens cannot be separated after collection, the
storage either at þ228C or þ88C up to the 6 h can be
allowed since most concentrations of analytes were suffi-
ciently stable when compared with those immediately
separated from cells. The stability of potassium, however,
is temperature-dependent and cannot be measured from
refridgerated blood specimen.
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