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Table 1 Major sources of variability in biochemical markers of bone turnover

From: The clinical utility of bone marker measurements in osteoporosis

Bone marker (Abbreviation) Source Action Advantages Disadvantages Analysis and sample type
Formation markers
Bone Alkaline Phosphatase (BAP) Enzyme present in osteoblast plasma membranes Enzymatic degradation of the mineralisation inhibitor pyrophosphate at alkaline pH Low intra-individual variability <10% [15] Not affected by renal function [15] Food has little effect [16] Long circulatory half-life 1–2 days [17] Sample stability [18] Cheap Up to 20% cross reactivity with liver isoforms [14] Changes with therapy minimal i.e. less than LSC of 25% [15] 2 peaks at 14:00 and 23:30 hrs Nadir 30% ↓at 06:30 [19] Multiple methodologies, can measure mass or activity [20] Automated and manual immunoassays Serum, EDTA plasma
Osteocalcin (OC) Major non-collagen bone Gla protein. Produced by osteoblasts during bone formation and bound to hydroxyapatite Influences osteoid mineralisation Provides negative feedback during remodelling process EDTA sample more stable [21] Late marker of osteoblast activity [15] Intact molecule unstable [15] Large inter-lab variation [20] Released during formation and resorption [17] Short half-life of a few minutes [22, 23] Influenced by Vit K status, renal function and circadian variability [15, 17] OC gene regulated at transcriptional level by 1,25-OH2 Vit D Vit K essential co-factor for γ-carboxylation of OC resulting in ↑ affinity for Ca and hydroxyapatite [14] Automated and manual immunoassays Multiplex microarray Serum, EDTA plasma
Procollagen type 1 Carboxy-terminal Propeptide (P1CP) Specific product of proliferating osteoblasts and fibroblasts. Cleaved from type 1 pro-collagen by proteases during type 1 collagen formation Quantitative measure of newly formed type 1 collagen Thermostability [14] Short half-life 6-8mins [14] Cleared in liver by mannose receptor so sensitive to thyroid hormones and IGF-1 [20] Highest levels 01:30 – 04:30, up to 20% higher than nadir 11:00 – 15:00 [19] Lacks sensitivity to changes during menopause [14] Automated and manual immunoassays Serum, EDTA plasma
*Procollagen type 1 amiNo-terminal Propeptide (P1NP) Specific product of proliferating osteoblasts and fibroblasts. Cleaved from type 1 pro-collagen by proteases during type 1 collagen formation Low intra-individual variability[20] Small circadian rhythm[15] Stable at room temp[21] Good assay precision[20] Superior for PMO monitoring - change from baseline ↑up to 80% with anti-resorptive and ↓up to 200% with PTH medication within 3months[15] Total assay affected by delayed clearance of monomeric fraction e.g. in renal failure or metastatic bone disease[24] Expensive Automated and manual immunoassays Multiplex microarray Total or Intact fractions Serum, EDTA plasma
Resorption markers Collagen derived
*Carboxy-Terminal cross-linked telopeptides of type 1 collagen (CTX) Type 1 collagen mainly bone Isomerisation to β aspartyl occurs in mature collagen Cleaved from type 1 collagen by cathepsin-K during bone resorption Variability↓ fasting[25] Sample stability, especially EDTA[18][21] Substantial ↓ post anti-resorptive treatment[26] Blood sample now preferential Large circadian variation – highest 01:30 – 04:30 approx 2x nadir 11:00–15:00[27] Automated and manual immunoassays Multiplex microarray Urine, serum, EDTA plasma
Carboxy-Terminal cross-linked telopeptides of type 1 collagen (ICTP or CTX-MMP) Newly synthesised type 1 collagen predominantly bone Cleaved from type 1 collagen by MMP during bone resorption   Large circadian variation [20] Influenced by renal and liver function [20] Not responsive to usual osteoporotic treatments [20] Manual immunoassay
amiNo-Terminal cross-linked telopeptides of type 1 collagen (NTX) Type 1 collagen mainly bone Cleaved from type 1 collagen by cathepsin-K during bone resorption Urine sample stable [15] uNTX sig predictor of fracture risk in postmenopausal women [28] Small dietary influence, although fasting blood sample preferred [15] Large circadian variation Influenced by renal and liver function [20] Units based on manufacturer’s calibrator i.e. bone collagen equivalents [15] Automated and manual immunoassays Urine, serum, EDTA plasma
Type 1 collagen alpha 1 helicoidal peptide (HELP) Type 1 collagen Amino acid 620–633 sequence of the α chain Cleaved from helical region of type 1 collagen by cathepsin-K during bone resorption High correlation to other markers of collagen degradation [14] 24 hr collection – hard to collect 2nd morning void with creatinine correction – additional analytical variability Clinical validity needs further investigation Manual immunoassay Urinary marker
Deoxypyridinoline (DPD) Mature type 1 collagen Cross link released when mature type 1 collagen breaks down Mechanically stabilise the molecule Reflect degradation of mature collagen only Specific to bone [14] Independent of dietary sources [20] Less invasive than blood 24 hr collection – hard to collect 2nd morning void with creatinine correction – additional analytical variability Circadian variation [20] Automated and manual immunoassays Urinary marker
Pyridinoline (PYD) Mature type 1 and 11 collagen Cross link released when mature collagen type 1 and 11 breaks down Mechanically stabilise the molecule Reflect degradation of mature collagen only Independent of dietary sources [20] Non-specific 24 hr collection – hard to collect 2nd morning void with creatinine correction – additional analytical variability Circadian variation [20] Influenced by liver function [20] Automated and manual immunoassays Urinary marker
Resorption markers Osteoclastic Enzymes
Tartrate Resistant Acid Phosphatase –isoform 5b (TRAP5b) Isoform of acid phosphatase, resistant to tartrate, cleaved by proteases into 5b, present in ruffled border of osteoclasts Cleaves type 1 collagen into fragments Characteristic of osteoclastic activity [14] Unstable at room temperature [22, 23] Circadian variability ↑ immediately after exercise [29] Automated and manual immunoassays Serum
Cathepsin K Cysteine protease present in ruffled border of actively resorbing osteoclasts Cleaves telopeptide and helical regions of type 1 collagen Specific biomarker of osteoclastic activity [14] Unstable at room temp Clinical validity needs further investigation Manual immunoassay Serum, EDTA plasma
Osteocyte activity markers
Receptor Activator of Nuclear factor Kappa B Ligand (RANKL) Produced by osteoblasts, activated by B and T cells Binds to RANK, which is expressed on osteoclasts and their precursors, stimulating their differentiation and activity Novel biomarker Provide safety, efficacy and pharmacokinetics data to confirm drug mechanisms and mode of action for future use Analytical problems Can measure free or OPG-bound [30] Circulating levels may not reflect bone microenvironment [31] Affected by thyroid function [32] Research method only Clinical and analytical validity needs further investigation Manual research –grade immunoassay Total or soluble forms in serum
Osteoprotegerin (OPG) Secreted by osteoblasts Decoy receptor to RANKL reduces bone resorption by binding to RANK and preventing osteoclastogenesis Novel biomarker Provide safety, efficacy and pharmacokinetics data to confirm drug mechanisms and mode of action for future use Affected by thyroid function [32] Research method only Clinical and analytical validity needs further investigation Manual research-grade immunoassay Serum
Dickkopf-related protein 1 (DKK1) Produced by osteocytes Inhibition of Wnt signalling pathway through binding to LRP5/6, blocking the Wnt effects on osteoblasts and decreasing bone formation Key role in regulation of bone turnover Research method only Clinical and analytical validity needs further investigation Manual research –grade immunoassay Serum
Sclerostin (SCL) Secreted by osteocytes Inhibition of Wnt signalling pathway through binding to LRP5/6, blocking the Wnt effects on osteoblasts and decreasing bone formation Significant ↓ with PTH therapy [33] Research method only Affected by immobility [34] ↑ in type 1 and 2 diabetes [35, 36] Clinical and analytical validity needs further investigation Manual research-grade immunoassaySerum
  1. *P1NP and CTX (highlighted in bold) are the markers of choice, recommended by the IOF, IFCC (2011) and NBHA (2012).
  2. ALP alkaline phosphatase, BSAP bone specific alkaline phosphatase, CTX carboxy-terminal cross-linked telopeptides of type 1 collagen, DKK-1 dickkopf-related protein 1, DPD deoxypyridinoline, EDTA ethylenediaminetetraacetic acid, HELP Type 1 collagen alpha 1 helicoidal peptide, ICTP carboxy-terminal cross-linked telopeptide of type 1 collagen, IGF-1 insulin-like growth factor-1, LRP low-density lipoprotein receptor-related protein, MMP matrix metalloproteinases, NTX amino-terminal cross-linked telopeptide of type 1 collagen, OPG osteoprotegerin, OC osteocalcin, PICP procollagen type 1 carboxy-terminal propeptide, PINP procollagen type 1 amino-terminal propeptide, PYD pyridinoline, RANK receptor activator of nuclear factor kappa B, RANKL receptor activator of nuclear factor kappa B ligand, SCL sclerostin, TRAP tartrate resistant acid phosphatase.