Myoglobin detection

Aslan, K. and Geddes, C. D. (2006). Microwave Accelerated and Metal Enhanced Fluorescence Myoglobin Detection on Silvered Sur ces Potential Application to Myocardial Infarction Diagnosis Plasmonics 1 53-59. 

Elevations in cTnl and cTnT are highly specific for myocardial injury. However, in individuals without myocardial disease, their levels are very low to undetectable. This is in contrast to the low but measurable concentrations of CK-2 and myoglobin detected in serum from skeletal muscle turnover in patients with noncardiac-related diseases and in normal individuals. Therefore release of cTnl or cTnT from myocardium into the blood following AMI and after the washout that accompanies successful reperfusion generates an excellent signal compared with no detectable baseline levels before myocardial damage. The initial rapid release of cardiac troponin subunits I and T following successful reperfusion is most hkely derived from the soluble cytosolic myocardial fraction (6% cTnT 3% cTnl). 

Cardiac myoglobin detection was based on direct electron transfer between the Fe(III)-heme and the electrode surface that was modified with metal nanoparticles stabilized by didodecyldimethylammonium bromide and antibodies. Gold, silver, and copper nanoparticles were tested as catalysts of the Fe(III)/Fe(II) electrode process. Experiments were carried out with human blood plasma samples of healthy donors and patients with acute myocardial infarction. The method proposed does not require labeled secondary antibodies. The myoglobin immunosensor has a detection limit of 5 ngrnL and a broad range of working concentrations (Suprun et al., 2011). The whole procedure takes 20 min and can be used to establish the diagnosis of acute myocardial infarction. 

Suprun, E.V., Shilovskaya, A.L., Lisitsa, A.V., Buiko, T.V., Shumyantseva, V.V., Archakor, A.I., 2011. Electrochemical immunosensor based on metal nanoparticles for cardiac myoglobin detection in human blood plasma. Electroanalysis 23 (5), 1051—1057. 

PSS-SG composite film was tested for sorption of heme proteins hemoglobin (Hb) and myoglobin (Mb). The peroxidaze activity of adsorbed proteins were studied and evaluated by optical and voltammetric methods. Mb-PSS-SG film on PG electrode was shown to be perspective for detection of dissolved oxygen and hydrogen peroxide by voltammetry with linear calibration in the range 2-30 p.M, and detection limit -1.5 p.M. Obtained composite films can be modified by different types of biological active compounds which is important for the development of sensitive elements of biosensors. 

FIGURE 4.47 Dependence of HETP on sample volume. Column Toyopearl HW-55F, 22 mm X 30 cm. Sample 0.1% myoglobin. Elution 14 mM Tris-HCI, pH 7.9, in 0.3 M NaCI. Flow rate 52 cm/ hr. Detection UV at 220 nm. Legend to. sample injection time Z, column length u. linear velocity. 

Fig. 4. HPHIC of standard proteins on the weak hydrophobic columns. The SynChro-pack PROPYL column was 25x0.41 cm Poly (alkyl aspartamid)-silicas were packed into 20 x 0.46 cm columns. Sample 25 pi containing 25 pg of each protein in buffer A. Buffer A 1.8 mol/1 ammonium sulphate + 0.1 mol/1 potassium phosphate, pH 7.0. Buffer B 0.1 mol/1 potassium phosphate, pH 7.0. Gradient 40-min linear 0-100% buffer B. Flow rate 1 ml/min. Detection A220 = 1-28 a.u.f.s. Peaks a = cytochrome C, b = ribonu-clease A, c = myoglobin, d = conalbumin, e = neochymotrypsin, / = a-chymotrypsin, g - a-chymotrypsinogen A [48]...

Following massive crush injury, myoglobin released from damaged muscle fibers colors the urine dark red. Myoglobin can be detected in plasma following a myocardial infarction, but assay of serum enzymes (see Chapter 7) provides a more sensitive index of myocardial injury. 

Figure 11.6 Positive ion electrospray mass spectra of an equimolar mixture of five standard proteins, under different instrumental settings, showing cases where prominent signals for the different charge states of (A) insulin, (B) ubiquitin, (C) cytochrome c, (D) lysozyme, and (E) myoglobin were preferentially observed, and (F) where signals for all the proteins were more uniformly detected.

FIGURE 16.8 HPLC chromatogram of cytochrome c and myoglobin digest, using a 250 cm x 4.6 mm ODS C18 Vydac column and a linear mobile-phase gradient, 5-50% B, in 50 min. Buffer A was 0.1% TFA in water and buffer B was 0.1 TFA in acetonitrile. UV detection was carried out at 214 nm, at room temperature (reprinted with permission from Electrophoresis). 

This same group developed a novel biosensor based on manganese myoglobin to detect nitric oxide. Mn-containing myoglobin was chosen as... 

S. Kroning, F.W. Scheller, U. Wollenberger, and F. Lisdat, Myoglobin-clay electrode for nitric oxide (NO) detection in solution. Electroanalysis 16, 253—259 (2004). 

Fig. 5. Effect of the flow rate on the separation efficiency. Separation of a protein mixture at six different flow rates (40,80,120,160,200 and 240 ml/min) normalized to the elution volume. Conditions Column 80 ml CIM DEAE Tube Monolithic Column Mobile phase buffer A 20 mM Tris-HCl buffer, pH 7.4 buffer B 20 mM Tris-HCl buffer + 1 M NaCl, pH 7.4 Gradient 0-100% buffer B in 200 ml Sample 2 mg/ml of myoglobin (peak 1), 6 mg/ml of conalbumin (peak 2) and 8 mg/ml of soybean trypsin inhibitor (peak 3) dissolved in buffer A Injection volume 1 ml Detection UV at 280 nm. (Reprinted with permission from Podgornik A, Barut M, Strancar A, Josic D, Koloini T (2000) Anal Chem 72 5693)...

Figure 3.23 Selectivity of phenyl and alkyl bonded stationary phase materials for protein separation. Column A, TSK gel phenyl-5PW RP, 75 mm x 4.6 mm i.d. B, TSK gel TMS 250, 75 mm x 4.6 mm i.d. eluent, 60 min linear gradient elution from 5% of 0.05% trifluoroacetic acid in 5%> aqueous acetonitrile to 80% of 0.05% trifluoroacetic acid in 80% aqueous acetonitrile flow rate, lml min-1 detection, UV 220 nm. Peaks 1, ribonuclease 2, insulin-, 3, cytochrome c 4, lysozyme-, 5, transferrin-, 6, bovine serum albumin-, 1, myoglobin-, and 8, ovalbumin.

Figure 4.8 Cation-exchange liquid chromatography of basic proteins. Column, Asahipak ES502C eluent, 20 min linear gradient of sodium chloride from 0 to 500 mM in 50 mM sodium phosphate buffer pH 7.0 flow rate, 1 ml min-1 temperature, 30 °C detection, UV 280 nm. Peaks 1, myoglobin from horse skeletal muscle (Mr 17 500, pi 6.8-7.3) 2, ribonuclease from bovine pancreas (Mr 13 700, pi 9.5-9.6) 3, a-chymotrypsinogen A from bovine pancreas (Mr 257 000, pi 9.5) and 4, lysozyme from egg white (Mr 14 300, pi 11.0-11.4). (Reproduced by permission from Asahikasei data)...

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