Large-gel two-dimensional electrophoresis-matrix assisted laser desorption/ionization-time of flight-mass spectrometry: an analytical challenge for studying complex protein mixtures.

The large-gel two-dimensional electrophoresis (2-DE) technique, developed by Klose and co-workers over the past 25 years, provides the resolving power necessary to separate crude proteome extracts of higher eukaryotes. Matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) provides the sample throughput necessary to identify thousands of different protein species in an adequate time period. Spot excision, in situ proteolysis, and extraction of the cleavage products from the gel matrix, peptide purification and concentration as well as the mass spectrometric sample preparation are the crucial steps that interface the two analytical techniques. Today, these routines and not the mass spectrometric instrumentation determine how many protein digests can be analyzed per day per instrument. The present paper focuses on this analytical interface and reports on an integrated protocol and technology developed in our laboratory. Automated identification of proteins in sequence databases by mass spectrometric peptide mapping requires a powerful search engine that makes full use of the information contained in the experimental data, and scores the search results accordingly. This challenge is heading a second part of the paper.

Alpha-cyano-4-hydroxycinnamic acid affinity sample preparation. A protocol for MALDI-MS peptide analysis in proteomics.

We present a new MALD1 sample preparation technique for peptide analysis using the matrix alpha-cyano-4-hydroxy-cinnamic acid (CHCA) and prestructured sample supports. The preparation integrates sample purification, based on the affinity of microcrystalline CHCA for peptides, thereby simplifying the analysis of crude peptide mixtures. Enzymatic digests can thus be prepared directly, without preceding purification. Prepared samples are homogeneous, facilitating automatic spectra acquisition. This method allows preparation of large numbers of samples with little effort and without the need for automation. These features make the described preparation suitable for cost-efficient high-throughput protein identification. Performance of the sample preparation is demonstrated with in situ proteolytic digests of human brain proteins separated by two-dimensional gel electrophoresis.

Aspergillosis with Aspergillus osteomyelitis and diskitis after heart transplantation: surgical and medical management.

This is the report of a 49-year-old man, who could be cured of aspergillosis with Aspergillus osteomyelitis and diskitis of the lumbar spine by medical and surgical therapy after heart retransplantation. Aspergillus osteomyelitis and diskitis is a rare complication of invasive aspergillosis. Lumbar involvement is the main manifestation, followed by rare locations in the tibia, ribs, wrist, sternum, pelvis, and knee. It is nearly always secondary to pulmonary aspergillosis, which occurs in about 6% to 14% of patients after heart transplantation. For treatment, a combined surgical and medical approach is recommended, which enables quick pain relief for the patient and histologic and microbiologic diagnosis and stabilization of the spine. Because of definitive cure of osteomyelitis cannot be guaranteed by removal of the abscess, medical therapy after surgery is necessary for years.

[A bipolar electrode with a radial asymmetrically aligned electric field. A new principle of nerve localization in regional anesthesia and pain therapy]. / Bipolare Elektrode mit radial asymmetrisch ausgerichtetem elektrischem Feld. Ein neues Prinzip der Nervenlokalisation in der Regionalaesthesie und der Schmerztherapie.

Electrostimulation is a common method of nerve localization. To improve the nerve-stimulating needle, we developed a new bipolar nerve-stimulating electrode with a more convenient extension of its electric field. Figure 1a shows the new nerve-stimulating needle; in Fig. 1b a longitudinal section is depicted. A hollow cannula (1) is surrounded by an isolator (5) that separates the internal electrode (1) from the external electrode (6). The surface of the latter is also covered by an isolator (8). Two cables, (2) and (7), connect the electrodes to a nerve stimulator. Syringes and tubes can be connected to the needle by using the attached part (4). Figs. 2a and b show top- and side views of the needle's sharp end. Except for small parts of the internal (1) and external electrode (2), both electrodes are covered by isolation (3). This arrangement leads to the radially asymmetric extension of the electric field. Studies with animal nerve-muscle preparations showed that the density of the electric field is higher, and thus electrostimulation can be carried out with up to 75% lower impulse amplitude as compared with insulated monopolar electrodes. The main advantage of the new arrangement of electrodes, however, is the dependency between revolution of the nerve-stimulating needle on its axis and the evoked muscle response. Muscle potentials measured by electromyography and respective degrees of needle torsion are shown in Fig. 3 (black circles: bipolar radially asymmetric electrode, white circles: insulated monopolar electrode).(ABSTRACT TRUNCATED AT 250 WORDS)

[Effect of spinal CSF dynamics on the subarachnoid diffusion of a substance applied close to the spinal cord]. / Der Einfluss der spinalen Liquordynamik auf die subarachnoidale Ausbreitung rückenmarksnah applizierter Substanzen.

In order to better understand the effects and side effects of intraspinal administration of morphine we studied the rostral spread of a comparable substance within the cerebrospinal fluid (CSF). This study was performed in connection with nuclear medical diagnostics ruling out possible rhinorrhoea or disturbances of CSF-circulation in 14 patients: Following lumbar intrathecal injection of the tracer 111-Indium-DTPA, the radioactivity over the medulla oblongata was measured continuously for 2 1/2 hours with a single probe scintillation counter; thereafter the distribution of activity over the total spinal canal was determined; finally the spread of activity was registered with the gamma scintillation camera in the 3rd, 24th and 48th hour. The diffusion of the tracer was followed in a model of the subarachnoid space. A few minutes after injection, activity over the medulla oblongata could be detected; initially it increased markedly, later less so; at the end of the 2 1/2 h observation time, approximately 8% of the total activity had reached this level. The timing of activity increase and the peak activity over the medulla oblongata varied between the individuals. Up to 48 hours the activity continued to shift from the spinal canal to the endocranium. Diffusion played a secondary role. These results are further evidence that morphine is transported cephalad within the CSF rather quickly and may act on cervical spinal cord and brainstem.

[Bupivacaine 0.75% versus 0.5% for peridural anesthesia. Sensory and motor blockade]. / Bupivacain 0,75% versus 0,5% zur Periduralanaesthesie. Sensible und motorische Blockade.

This study was designed to investigate under controlled conditions sensory and motor blockade provided by epidural anaesthesia following two concentrations of bupivacaine without adrenaline. Twenty four patients received for extracorporeal shock wave lithotripsy a lumbar continuous epidural anaesthesia randomized with bupivacaine 0.75% (n = 12) or 0.5% (n = 12). During development and regression, sensory blockade was determined by the pinprick method, motor blockade by the Bromage score. All differences between the two concentrations--even though not all statistically significant--spoke in favour of bupivacaine 0.75%: shorter time of onset, more cephalad spread, higher intensity and longer duration of sensory and motor blockade. The higher concentration should be injected more slowly due to the increased risk if intravascular injection should occur; it should not be used for cesarean section.

Epidural morphine: a clinical double-blind study of dosage.

The purpose of this randomized double-blind study was to determine the optimal dose of epidural morphine by establishing a dose-effect relationship. The 139 patients, who had orthopedic operations on the lower extremities, received continuous lumbar epidural anesthesia with bupivacaine, 0.75%, with or without the addition of 1, 2, 3, 4, or 5 mg of morphine hydrochloride. Analgesia and side effects were determined during the first 24 hr postoperatively. In the 12-hr period after epidural anesthesia, arterial blood gas tensions were compared between those patients who received 5 mg morphine (n = 13) and those who received no morphine (n = 14). Patients who received 2 or more mg of morphine were less likely to require the administration of postoperative systemic analgesics (P less than 0.05). The addition of 2 or more mg of morphine to bupivacaine, 0.75%, reduced postoperative pain intensity (P less than 0.05); 5 mg of morphine reduced pain intensity for the longest time. Frequency of catheterization and pruritus increased dose-dependently. The mean PaCO2 after 5 mg of epidural morphine averaged 5 mm Hg higher than in the control group, indicating minor respiratory depression, better analgesia, or both. The dose of 3 mg of epidural morphine added to the local anesthetic is recommended for postoperative analgesia after surgery of the lower extremity; it is a compromise that provides adequate analgesia with an acceptably low frequency and intensity of side effects.

[0.5 mg intrathecal morphine in spinal anesthesia. A double blind study on sensory block, postoperative analgesia and adverse effects]. / ,5 mg Morphin intrathekal bei Spinalanaesthesie. Eine Doppelblindstudie zu sensibler Blockade, postoperativer Analgesie und Nebenwirkungen.

In a randomized double-blind study the influence of morphine 0.5 mg on the development and regression of spinal anaesthesia, the postoperative analgesia and the side effects were investigated. Forty-two patients received an isobaric spinal anaesthesia with tetracaine 20 mg without morphine (n = 19) or with morphine 0.5 mg (n = 23). The sensory blockade was tested by pinprick; the patients evaluated their postoperative pain with an analogue scale. Arterial blood gases, respiratory rate, blood pressure and heart rate were measured and side effects determined. In the test group the cranial level of anaesthesia was during the development (p greater than 0.05) and regression (p less than 0.05) half to three segments higher than in the control group. The postoperative analgesia was more intense and longer lasting with morphine than without (p less than 0.05). Following morphine, P art CO2 was higher (p less than 0.05), the respiratory rate lower (p less than 0.05). Pruritus, nausea, vomiting and disturbances of micturition were more frequent. Following spinal anaesthesia with a deeper level of anaesthesia at T8-T11 the postoperative analgesia was superior than following spinal anaesthesia with a higher level of anaesthesia at T3-T4 (p less than 0.05). Only following higher levels of anaesthesia there was evidence of respiratory depression (p less than 0.05). This is why the level of spinal anaesthesia with the addition of morphine must not be higher than necessary for surgery.

Epidural buprenorphine--a double-blind study of postoperative analgesia and side effects.

Epidural buprenorphine was investigated as a postoperative analgesic in a randomized double-blind study of 158 patients given epidural analgesia with mepivacaine or bupivacaine for orthopedic surgery of the lower extremity. At the end of surgery, patients were given either 0.15 mg of epidural buprenorphine (n = 38), 0.3 mg (n = 37) in 15-ml saline, or no further epidural injections (n = 47, control group) after 2% mepivacaine for intraoperative anesthesia. A fourth group (n = 36) received 0.3 mg of buprenorphine in 15-ml saline, after the intraoperative use of 0.5% bupivacaine. The patients rated postoperative pain. The need for additional analgesics as well as side effects were recorded. Analgesia after 0.15 mg buprenorphine was superior to that after no reinjection for 6 hr after surgery (P less than 0.05). Buprenorphine (0.3 mg) was superior both to no reinjection and to 0.15 mg of buprenorphine until the twelfth hour (P less than 0.05). Analgesia after bupivacaine followed by 0.3 mg of buprenorphine was not significantly different than analgesia seen after mepivacaine followed by 0.3 mg of buprenorphine. There was an increase of PaCO2 of 2-5 mm Hg between 1.5-3.5 hr after 0.3 mg of buprenorphine without any evidence for late respiratory depression. Other side effects, e.g., disturbances of micturition, pruritus, nausea, vomiting, fatigue, and headache, were comparably common in all groups. The epidural administration of 0.3 mg buprenorphine may be recommended for postoperative analgesia following orthopedic surgery of the lower extremity.

[Peridural anesthesia with etidocaine. Clinical studies on the effect of vasoconstrictors on sensory and motor blockade]. / Periduralanaesthesie mit Etidocain. Klinische Untersuchungen zum Einfluss von Vasokonstriktoren auf die sensible und motorische Blockade.

In a randomized double-blind study etidocaine 1.5% without a vasoconstrictor (n = 10), with adrenaline 1:200,000 (n = 10) and with ornipressine 1 IU/10 ml (n = 10) were administered epidurally to 30 orthopaedic patients. Sensory blockade was tested with electric pain stimuli; motor blockade was assessed with dynamometry during isometric plantar flexion of the foot and with the Bromage score; the intraoperative analgesia was determined as well. The development of sensory and motor blockade became faster and more intensive by addition of the two vasoconstrictors. The intraoperative analgesia, insufficient without vasoconstrictors, however, not to full satisfaction. Adrenaline improved the effects of etidocaine more than did ornipressine . The sole use of etidocaine without and with vasoconstrictors is not recommended for epidural anaesthesia for surgery of the lower extremities. It is the local anaesthetic of choice for intensifying motor blockade during already achieved analgesia.

Assessment of motor blockade during epidural anesthesia.

Motor blockade during epidural anesthesia was assessed using dynamometry to measure the maximum force of plantar flexion, and the Bromage score method. In a randomized double-blind study, 40 orthopedic patients received one of the following four local anesthetics epidurally: 0.5% bupivacaine-HCl (n = 10), 0.5% bupivacaine-CO2 (n = 10), 1.0% etidocaine (n = 10), and 1.5% etidocaine (n = 10), all without addition of a vasoconstrictor. The decrease of force, measured by dynamometry, followed an e function. This function permits the description of motor blockade development using one single number, its half-life. This finding corresponds to the increase of motor nerve fiber blockade due to diffusion of the local anesthetic into the nerve. Dynamometry was more precise than the Bromage score in describing motor blockade. Both the Bromage score and dynamometry produced roughly comparable results of onset, rate of development, and maximum intensity of motor blockade. These parameters of motor blockade were superior after both etidocaine solutions compared to both bupivacaine solutions. Neither carbonization of bupivacaine nor increased concentration of etidocaine significantly improved motor blockade produced by these drugs. Dynamometry is a time consuming and costly method for accurate quantification of motor blockade during epidural anesthesia; the Bromage score is more practical under clinical considerations and provides useful information.

The extent of blockade following various techniques of brachial plexus block.

The extent of sensory and motor blockades was examined in 195 patients 5 and 20 min after four different techniques of brachial plexus block using 50 ml of 0.5% bupivacaine. The interscalene technique of Winnie (N = 50) resulted in a preferential blockade of the caudad portions of the cervical plexus and the cephalad portions of the brachial plexus. The supraclavicular approach of Kulenkampff (N = 55) and the subclavian perivascular approach of Winnie (N = 56) each resulted in a homogeneous blockade of the nerves of the brachial plexus. The Winnie modification of the axillary approach (N = 34) resulted in a preferential blockade of the caudad nerves of the brachial plexus. With all four techniques, motor blockade developed faster than sensory blockade. The difference in results suggests that the approach to be used should depend primarily upon the site of the operation. The perineural space enclosing the brachial plexus greatly facilitates the spread of a local anesthetic when injected; however, it is usually not filled completely or evenly.

Epidural morphine for postoperative analgesia: a double-blind study.

Postoperative analgesia and the side effects of epidurally injected morphine were investigated in a double-blind study. Following lumbar epidural anesthesia for orthopedic operations, 174 patients received, in a randomized, double-blind fashion, either 0.1 mg/kg of morphine epidurally, 0.1 mg/kg of morphine intramuscularly, or saline epidurally at the end of surgery. Following epidural morphine, postoperative pain was les frequent, less intense and of shorter duration, use of analgesics and sedative was less frequent; and the postoperative feeling of well-being rated better than after systemic morphine or epidural saline. These effects were more frequent when bupivacaine was used for operative epidural anesthesia than when mepivacaine was used. The results were age independent. Side effects following epidural morphine included pruritus and disturbances of micturition. Nausea, vomiting, fatigue, and headache were of comparable frequency in the three groups.

[Model investigations regarding the spread of local anaesthetics in isobaric spinal anaesthesia (author's transl)]. / Modelluntersuchungen zur Ausbreitung der "isobaren" Spinalanaesthesie.

The better to comprehend isobaric spinal anaesthesia, we constructed a thermoregulated model of the subarachnoid space. With it we investigated the six not exactly isobaric local anaesthetics bupivacaine, carticaine, lidocaine, mepivacaine, prilocaine and tetracaine, in order to determine the effect of the following factors on their spread: injected volume, position, density, temperature of the injected solution, rate of injection, barbotage, direction of the spinal needle and its opening. The injected volume was most important. The small differences in density were of consequence in the vertically mounted model. Higher rates of injection increased the spread. The remaining factors were not relevant. The investigation corroborates the clinical observation that spinal anaesthesia with approximately isobaric solutions is a well controllable technique.

[Density of cerebrospinal fluid and local anesthetics (author's transl)]. / Dichte von Liquor und Lokalanaesthetika.

The densities of cerebrospinal fluid and of local anesthetics, applicable to isobaric spinal anesthesia, were determined by using the Digital Density Meter DMA 02. The density of CSF showed little variation and at 37 degrees C was 1.00021 +/- 0.00024 g/cm3 (mean +/- SD, n = 22). The density of the local anesthetics bupivacaine 0.5%, carticaine 2%, lidocaine 2%, mepivacaine 2% and prilocaine 2% varied at 25 degrees C between 1.001 and 1.005 g/cm3, at 37 degrees C between 0.997 and 1.001 g/cm3. Tetracaine 0.5% in CBF increased its density by 0.00046 g/cm3. The addition of the vasoconstrictors adrenaline and ornipressine (POR 8) increased the density of the local anesthetic solutions insignificantly. On the basis of the narrow range of variation of CSF density, reliable statements may be made on the density dependent spread of local anesthetics in spinal anesthesia.

[Evaluation of brachial plexus block. Comparison between supraclavicular and interscalene approach (author's transl)]. / Beurteilung der Plexus brachialis-Blockade. Vergleich des supraklavikulären und interscalenären Zugangs.

A scheme for evaluating brachial plexus block was developed, which is based on anatomical structures and which utilizes simple neurological examination techniques. It facilitates the localization of the tip of the cannula near the plexus, gives an idea of the spread of the local anesthetic in the region of the trunks and cords, gives well timed judgement on the success of the block and allows a comparison of the development of the block when using various techniques. Using this scheme, the development of the block was examined following use of the supraclavicular and interscalene approach. With the supraclavicular technique, motor as well as sensory blockade of all nerves of the brachial plexus occurred with about the same frequency; with the interscalene technique, the centre of the block affected the caudal nerves of the cervical plexus and the cranial nerves of the brachial plexus. Following both approaches, the blockade developed from proximal to distal areas, the motor blockade preceding the sensory blockade.

[Changes of carbohydrate and fat metabolism under spinal anaesthesia or neuroleptanaesthesia and operation (author's transl)]. / Veränderungen im Kohlenhydrat- und Fettstoffwechsel unter Spinal- bzw. Neuroleptanaesthesie und Operation.

The effects of spinal anaesthesia neuroleptanaesthesia and surgery on the metabolism of carbohydrates and fats were investigated in two groups of 11 young, metabolically healthy patients, undergoing trauma surgical procedures. For both groups the determined parameters glucose, lactate, glycerol, ketone-bodies, NEFA and insulin remained, during the operative and early postoperative phase, within the physiological range without any marked differences. Both anaesthetic procedures seemed to be equally suited for these operations; therefore other criteria should be used as a basis for the selection of the anaesthetic management.

[Isobaric spinal anaesthesia with bupivacaine and tetracaine (author's transl)]. / Isobare Spinalanaesthesie mit Bupivacain und Tetracain.

4 ml 0.5% solutions of bupivacaine and tetracaine without the addition of a vasoconstrictor, approximately isobaric, were used for spinal anaesthesia on patients in the sitting position. The sensory and motor block due to the two local anesthetics was tested and compared. The mean time on onset of complete analgesia was the same for both local anaesthetics (9 and 11 min), as was also the highest level of analgesia (T10). The duration of maximal extension of analgesia was on an average 45 min longer due to tetracaine (bupivacaine 105 min, tetracaine 150 min). The duration of maximal spread of the blocked sensation of pain, temperature, pressure and touch was similar for each of both local anesthetics. The regression of these sensory qualities, blocked in a dissoaciated manner, took a parallel course. With tetracaine the motor block of the lower extremities developed faster and lasted longer (Bromage 3 for bupivacaine 192 min, for tetracaine 220 min). Motor function and proprioception normalized in a synchronized manner. Isobaric spinal anaesthesia with these two solutions of local anaesthetics was found to be reliable and controllable, especially when administered to the sitting patient. Tetracaine is a good alternative to bupivacaine, currently controversial for intrathecal use.

[An improved bronchoscope (author's transl)]. / Ein verbessertes Beatmungsbronchoskop.

This bronchoscope differs from those normally used in the following ways: 1. In the endobronchial position the bronchoscope makes possible improvement in ventilation of the contralateral lung by increasing the area of the side-openings above the bifurcation. - 2. An inflatable cuff on the outer tube seals the bronchoscope against the trachea like a breathing tube. - 3. Jet-ventilation van be employed when the bronchoscope is open.

[Advantages of blood gas analyses of an improved ventilation bronchoscope (author's transl)]. / Blutgasanalytisch nachgewiesene Vorteile eines neuen Beatmungsbronchoskopes.

During bronchoscopy in elder patients the development of respiratory acidosis is a hazard. A new and improved ventilation bronchoscope decreases the risk, which was demonstrated by blood gas analyses. This bronchoscope differs from those normally used in the following ways: 1. An inflatable cuff on the outer tube seals the bronchoscope like a breathing tube against the trachea. 2. This bronchoscope, although in the endobronchial position, improves the ventilation of the contraleateral lung aided by wider side holes lying above the bifurcation. 3. Jet-ventilation can be employed.