Homemade Glove Retrieval Bag in Laparoscopic Complete Excision of Benign Ovarian Teratoma with Ovarian Tissue Preservation in Children: A Case Series.

STUDY OBJECTIVE: To present our experience of laparoscopic resection of pediatric benign ovarian teratomas with gonadal preservation, using a homemade glove retrieval bag. DESIGN, SETTING, PARTICIPANTS, INTERVENTIONS, AND MAIN OUTCOME MEASURES: Review of all girls with benign ovarian teratomas who were managed with laparoscopic ovarian-sparing surgery (OSS) at our hospital between January 2013 and December 2018. RESULTS: Eleven patients were included for analysis with a mean age of 6.1 years. Ten patients received elective surgery, whereas 1 patient received emergency surgery because of ovarian torsion. Main indication for OSS was the existence of a dissection plane between tumor margins and healthy ovarian tissue. Postoperative outcome and follow-up were uneventful with a median follow-up of 30.1 months (range; 12-60 months). CONCLUSION: Laparoscopic OSS can be safely performed for these tumors. Laparoscopic magnification with energy devices are excellent tools in such procedures. The homemade glove bag can be used to retrieve the tumor effectively in countries with limited resources.

Rapid tissue processing using a temperature-controlled collection device to preserve tumor biomarkers.

Precision tissue diagnostics rely on high quality input specimens so that assay results are not affected by artifact, but advances in collection and processing of tissue specimens have lagged behind innovations in diagnostic assay development. Therefore, we have designed and evaluated a novel surgical tissue collection device that maintains and monitors sample temperature and motion throughout transport so that the major preanalytical variable of tissue temperature can be controlled and measured. This device, in combination with an improved cold-hot tissue fixation protocol affords optimal biomarker preservation in less overall time, thereby simultaneously improving diagnostic quality and turnaround time. We collected 50 primary and metastatic liver tumors using a novel transport device. Tissue was fixed using a rapid cold-hot fixation protocol and immunohistochemical assays were used to assess the performance of the device, in comparison to control tissue preserved using standard clinical fixation protocol. Two pathologists evaluated the IHC studies in a blinded fashion to determine the immunophenotype of each tumor. The observed IHC staining intensities and the clinical impressions of the immunophenotypes did not differ between tissue collected with the novel device and control tissue, while improvements in processing time were achieved. The novel cold transport device and rapid fixation protocol can be successfully and safely combined and used to monitor specimen conditions, thus preserving the diagnostic utility of specimens and improving the overall turn-around time of the diagnostic process.

Tissue preservation through socket-shield technique and platelet-rich fibrin in immediate implant placement: A case report.

RATIONALE: In this report, a combination of socket-shield technique (SST) and platelet-rich fibrin (PRF) technique was used for immediate implant placement on a fractured central incisor. During the follow-up visit, cone beam computed tomography (CBCT) and clinical observation were used to evaluate the preservation outcome of peri-implant bone and gingiva. PATIENT CONCERNS: The patient was a 28-year-old healthy female patient who desired her fractured 21 to be replaced with an implant-supported single crown; the fractured 21 comprised a post-core crown with insufficient residual bone at the labial site. DIAGNOSIS: The root of 21 exhibited a complex root fracture; the labial portion of the alveolar ridge was thin (<1 mm) and partial ankylosis of the residual root was observed. INTERVENTIONS: Modified SST was applied to the labial portion of the residual root. The implant was placed immediately at the lingual site of the retained socket-shield root fragment; PRF was the placed in the gap between the root fragment and the implant. Final prosthodontic treatment was performed at 24 weeks after implant placement. OUTCOMES: Clinical examination and CBCT scanning at various follow-up visits time showed that the periodontal tissue was well- preserved. At 6 months after surgery, the average horizontal and vertical peri-implant bone resorption was 0.4 mm; a follow-up visit at 18 months post-loading indicated that peri-implant tissue was well preserved by the shield-technique and no significant peri-implant tissue resorption was displayed. LESSON SUBSECTIONS: In cases of anterior teeth with intact but insufficient residual alveolar ridge, the SST with PRF may be effective for preservation and maintenance of stable peri-implant tissue.

A Mobile Extracorporeal Extremity Salvage System for Replantation and Transplantation.

BACKGROUND: Traumatic amputation is the second leading cause of limb loss in the United States. The preferred treatment is salvage and replantation of the amputated limb, whenever possible, and allotransplantation is a novel procedure whereby healthy limbs are procured from deceased organ donors and transplanted into the amputee recipient. A major restriction for both procedures is the irrecoverable muscle damage occurring due to ischemia. We investigated the feasibility of using a novel lightweight, mobile perfusion device specifically designed to perfuse amputated porcine limbs with an acellular perfusion solution to delay ischemic muscle damage prior to transplantation or replantation. METHODS: Bilateral hind limbs of Yorkshire pigs were amputated; one of the limbs was preserved by perfusion in the mobile perfusion device, and the other by storage in ice slurry for 12 hours. RESULTS: Five sets of bilateral limbs were preserved as described previously. A defined pressure of 30 mm Hg was reliably maintained in the arterial system without loss of flow. Comparison of the perfusate composition before and after limb passage revealed significant differences. Muscle biopsies showed a consistent progression of clusters of hypoxic cells in the control limbs with time. Similar changes could not be observed in the perfused tissue. CONCLUSIONS: We have designed and built a small, mobile perfusion device that is operational and that more closely mimics the normal physiological environment when compared with the current standard of preservation in ice slurry. This project may have far-reaching implications for the treatment of limb loss through replantation and transplantation.

[Midterm Outcomes of Root Replacement Using Homograft for Aortic Valve Infective Endocarditis].

From August 2003 to June 2013, 9 patients with aortic valve endocarditis underwent aortic root replacement using homografts which were harvested and preserved in our institute. The median patient age was 62 years (range 46~84) and 5 patients were men. Four cases were prosthetic valve infections. The in-hospital mortality was 0%. In 8 of 9 cases were evaluated on midterm outcomes. At a median of 52 months (range 19~156), overall survival was 100%, freedom from cardiovascular events was 87.5%. The peak aortic pressure gradient was 9.04 ± 4.2 mmHg. Aortic regurgitation was less than 2 of 4 in all cases.

Histologic validation of vacuum sealed, formalin-free tissue preservation, and transport system.

We describe five validation trials of new vacuum sealing technologies that change the approach to the preanalytic "front end" of specimen transport, handling, and processing and illustrate their adaptation and integration into existing Lean laboratory operations with reduction in formalin use and personnel exposure to this toxic and potentially carcinogenic fixative. These trials provide histologic assessment by numerous pathologists of tissues processed in this new paradigm and define the financial advantages of applying this technology to the postanalytic or "back end" process of tissue storage. We conclude that the TisssueSAFE and SealSAFE vacuum sealing systems are both promising technologies for preserving fresh human specimens that can promote a safer environment by markedly reducing formalin use in operating room theaters and can minimize formalin use by laboratories.

Towards the lean lab: the industry challenge.

In anatomic pathology, the current state encompassing the pre-analytic processes of tissue collection, handling, examination, preparation, processing, and storage are largely uncontrolled, inconsistently performed, and/or not standardized according to the sound scientific data. Pre-analytic defects result in nearly three-quarters of the problems in laboratory diagnostics. This is evident in quality surveys from well-respected institutions that document high miss rates in the required basics of information related to patient and tissue identity, let alone parameters documenting quality aspects related to the surgical specimen and its preservation. This talk will describe the historical approach to tissue processing and identify gaps from worldwide observations in current laboratory practices. It will also offer potential methodological and technological solutions and process improvements that laboratories may consider in serving the ultimate users of pathology information: the clinician and the patient. It illustrates the need for scientifically validated specimen guidelines and a performance based, standardized and documented "chain of custody" of the pre-analytical steps from the patient's body through fixation. For thought leaders and professional standard setters, opportunities for optimizing molecular studies exist in specimen collection, transfer, grossing, fixation, and decalcification protocols. In this evolving era of molecular profiling and personalized therapeutic decision-making, a well-reasoned and coordinated focus on pre-analytic processes that optimizes specimens for subsequent testing will result in: Improved specimen quality for molecular testing Improved accuracy of diagnostic and molecular test results Reduced Turnaroundtimes for same-day diagnosis Enhanced satisfaction of clinicians and patients.

The effect of time, temperature and storage device on umbilical cord blood gas and lactate measurement: a randomized controlled trial.

OBJECTIVE: Umbilical cord blood gas analysis has a significant and growing role in early neonatal assessment. Factors often delay analysis of cord blood allowing values to change. Consequently, this study evaluates the impact of time, temperature and method of storage on umbilical blood gas and lactate analyses. METHODS: Umbilical cord segments from 80 singleton deliveries were randomized to: cords at room temperature (CR), cords stored on ice (CI), syringes at room temperature (SR) or syringes stored on ice (SI). Analysis occurred every 15 minutes for one-hour. Mixed model analysis of variance allowing for repeated measures was utilized. RESULTS: Cord arterial pH deteriorated in CR, CI, and SI within 15 minutes (p ≤ 0.001), with SR stable until 60 minutes (p = 0.002). Arterial pCO(2) remained stable in SR and CI, increased in SI (p = 0.002; 45 minutes) and decreased in CR (p < 0.001; 45 minutes). Arterial base excess deteriorated in CR and SI (p ≤ 0.009; 15 minutes), SR (p < 0.001; 30 minutes), and CI (p < 0.001; 45 minutes). Arterial lactate levels increased within 15 minutes in all groups (p < 0.001). CONCLUSIONS: Cord blood gas values change rapidly after delivery. Smallest changes were seen in SR group. Data suggest that analyses should be conducted as soon as possible after delivery.

Scanning electron microscopy of cell surface morphology.

The surface of metazoan cells is a landscape not clearly visualized by light microscopy. Many cells elaborate protrusive structures such as microvilli, filopodia, lamellipodia, and surface ruffles that play important roles in the interaction between the cell and its environment. The high resolution of scanning electron microscopy makes it an ideal technique for studies of the cell surface; however, preservation of fine surface structure can be problematic. Here we highlight the critical factors in sample preparation to ensure optimal high-resolution imaging of the surface of mammalian cells and tissues.

Quality control of bacterial contamination in autologous peripheral blood stem cells for transplantation.

BACKGROUND AND OBJECTIVES: Microbiological follow-up is part of quality control of peripheral blood stem cell (PBSC) manipulation. DESIGN AND METHODS: We prospectively studied microbiological cultures performed in 865 consecutive untreated autologous PBSC harvests from 348 patients. Our aim was to know the rate of microbiological contamination, the optimum moment to evaluate the sample and the clinical significance of the positive findings. RESULTS: Fifty-nine of the 852 samples (6.9%) yielded a positive culture after PBSC collection (sample 1) and 62 samples also yielded positive results before cryopreservation (7.2%) (sample 2). At the time of the analysis, a total of 520 aphereses had been infused and the number of positive cultures after thawing (sample 3) and after washing (sample 4; 82 aphereses) was 5.4% and 2.3%, respectively. Most of the positive cultures were due to coagulase-negative staphylococci (48 isolates). After thawing 15 coagulase-negative staphylococci and 2 enterococci isolates were recovered. Comparison between samples using a marginal homogeneity test showed no differences in the rate of contamination observed at the different sampling points. INTERPRETATION AND CONCLUSIONS: Positive microbiological findings in collected PBSC are not due to contamination within the laboratory. Cryopreservation using DMSO does not eradicate bacteria and manipulation does not seem to affect results. To simplify the procedure it would be possible to eliminate the microbiological controls performed immediately before cryopreservation.

Preclinical evaluation of an automated closed fluid management device: Cytomate, for washing out DMSO from hematopoietic stem cell grafts after thawing.

Infusion of dimethylsulfoxide (DMSO) contained in cryopreserved and thawed hematopoietic stem cell (HSC) grafts is frequently associated with mild or moderate adverse reactions, and occasionally with more severe events including neurological symptoms. The severity of these complications is related to the amount of residual DMSO. We evaluated a recently available, closed, automated and 'cgmp (current good manufacturing practice) compliant' device (CytoMate) for its ability to wash out DMSO at the expense of a limited loss of viable CD34(+) cells. A total of 16 procedures were carried out with 39 blood HSC bags intended for destruction. Mean amounts of DMSO for each cellular product (one, two or three bags) were between 12.2 and 39.6 g before thawing; after the washing procedure, residual DMSO quantities were between 0.1 and 3.7 g. When set up to reproducibly allow for a more than 96% elimination of DMSO, processing of thawed cells with the CytoMate cell processor resulted in a mean recovery of viable total cells, CD34(+) cells and lymphocyte subsets above 60%. We conclude that this simple and efficient washing technique is suitable for routine processing of HSC grafts. Clinical studies will demonstrate whether a reduction in the incidence of adverse effects associated with DMSO infusion is observed.

[Significance of extracorporally perfused microsurgical free flaps]. / Zum Stellenwert freier extrakorporal perfundierter mikrochirurgischer Gewebelappen.

BACKGROUND: Since 30 years microvascular reconstruction has been established in reconstructive plastic head and neck surgery. Since 10 years an extracorporal perfusion set for the vital preservation of microsurgical free tissue flaps of several days has been developed. Now it is the question which significance the extracorporal perfusion of free flaps has today. METHOD: The spectrum of the actual utilization is the parameter which determines the importance. A recommendation on the safe period of use for the vital preservation of microsurgical free flaps making use of the extracorporal perfusion system in room temperature (21 degrees C) is to be developed on the porcine vertical musculocutaneous trapezius flap. RESULTS: The device is regularly used in experimental plastic surgery for the research of pathophysiology of microvascular free flaps. It is frequently used in biotechnology for pharmacological and toxicological tests of the resorption of drugs through the skin and it is useful for the microsurgical training of free flap transplantation. The minimal period of a safe function is 46 hours. CONCLUSIONS: The device is a useful test system for the research of the ischemic period of microsurgical free flaps. Its greatest benefit is its use as a biotechnical test system reducing at the same time the consumption of animals. It is also a useful training device in reconstructive head and neck surgery. It is not suitable for the clinical free flap transplantation in human patients.

Catastrophic failures of freezing bags for cellular therapy products: description, cause, and consequences.

BACKGROUND: Container integrity is critical for maintaining sterility of cryopreserved cellular therapy products. We investigated a series of catastrophic bag failures, first noticed in early 2001. METHODS: Process records were reviewed for all PBPC and lymphocyte products cryopreserved in bags from January 2000 through April 2002. Patient charts were also reviewed. RESULTS: One thousand two hundred and four bags were removed from storage for infusion to 261 patients. All products had been cryopreserved in Cryocyte poly(ethylene co-vinyl acetate) (EVA) bags in either 10% DMSO or 5% DMSO and 6% pentastarch. Product volumes were 25-75 mL, and bags were stored with overwrap bags in a liquid nitrogen tank. From January 2000 to April 2001, failure occurred in 10 of 599 (1.7%) bags. From May 2001 to April 2002, 58 of 605 (9.6%) bags failed, typically with extensive fractures that were visible before thaw. Of the 58 that failed, 24 were salvaged by aseptic methods and infused to patients under antibiotic coverage; 10 of those 24 (42%) had positive bacterial cultures. Bag failures were not related to product type, cryoprotectant solution, liquid versus vapor storage, or freezer location. Failures were linked to use of four Cryocyte bag lots manufactured in 2000 and 2001. After replacing these lots with a 1999 Cryocyte lot and with KryoSafe polyfluoroethylene polyfluoropropylene (FEP) bags, no more failures occurred in 75 and 102 bags, respectively, thawed through April 2002. DISCUSSION: High rates of bag failure were associated with four Cryocyte bag lots. No serious adverse patient effects occurred, but bag failures led to microbial contamination, increased product preparation time, increased antibiotic use, and increased resource expenditure to replace products.

Thawing of cryopreserved mobilized peripheral blood--comparison between waterbath and dry warming device.

BACKGROUND: Thawing of cryopreserved mobilized peripheral blood (MPB) is routinely performed for autologous and allogeneic MPB transplantation. Usually thawing is achieved by submerging the cell bag in a waterbath (37 degrees C temperature). We compared the effectiveness of thawing cryopreserved MPB in a waterbath with an electric dry-warming device containing warmed gel pads (Sahara, Transmed). METHODS: Two cryopreserved bags from each of 31 apheresis procedures were thawed in a waterbath and under dry conditions in parallel. Viability (dye exclusion), apoptosis/necrosis (annexin/propidiumiodide staining) and clonogenic potential (CFU-E plus BFU-E, CFU-GM) of the cells were tested after thawing. RESULTS: Statistical analysis by Wilcoxon matched-pair test showed no significant difference between the thawing procedures in terms of the in vitro parameters tested. DISCUSSION: Our results indicate that thawing of cryopreserved MPB using dry warming and water bath give similar viability, apoptosis/necrosis rate and clonogenic potential. Both procedures take about the same amount of time and are easy to perform. Nevertheless, the potentially decreased risk of bacterial contamination of either the cell product or the patient room, and guidelines of good clinical practice (GCP), favor the use of the dry warming procedure.

[An example for stepwise reduction of work site-induced formaldehyde exposure in pathology]. / Beispiel für eine schrittweise Verminderung der arbeitsplatzbedingten Formaldehydbelastung in der Pathologie.

Occupational formaldehyde exposure in pathology depends on the efficiency of the ventilation system in use and may reach concentrations considerably above the current threshold limit values. The reduction of formaldehyde exposure by stepwise improvement of the ventilation system at a pathologist's workplace is presented as an example in this paper. Assessment of formaldehyde concentration by personal air sampling at a workplace originally equipped with a hood ventilation system resulted in values of up to 4 ml/m3 in the pathologist's breathing zone. Lowering the ventilation inlet to the working level by connecting the hood to a suction unit via a flexible hose resulted in an effective reduction of formaldehyde exposure to values of about 0.5 ml/m3. This simple and low-cost technical improvement had some uncomfortable side effects, such as current noise and wind chill, which could only be overcome by installing ventilated work tables according to modern technical standards.

New dimensions for the vital storage of microsurgical free flaps: an experimental approach.

A revolutionary circulatory system has been developed to nourish big, free osteomusculocutaneous flaps extracorporally. Thus we will be able to transplant the free flaps to defect areas that have no sufficient vascular situation. In respect to the cold storage of microsurgical free flaps, to date maximal periods of ischaemic tolerance have been considerably exceeded; the maximal period is currently 168 hours. The vitality of the flap is monitored through parameters setting forth the consumption of oxygen together with histology and electron microscopy. The oxygenation of the nutritive medium is achieved through an aqueous phase gas exchange.

Autologous transplant programs in community hospitals: role of centralized stem cell processing laboratories.

Transplantation of bone marrow and peripheral blood stem cells was initially limited to large, research-oriented institutions. General acceptance of autologous transplantation has grown rapidly in recent years and there is now interest in the establishment of autologous transplant programs in community hospitals. However, the lack of facilities to process and cryopreserve stem cells can be a major obstacle to small transplant programs. In response to this problem, we suggest that a centralized stem cell processing facility can offer the required services to several hospitals within a metropolitan area. The establishment of processing facilities within currently existing blood banks and blood centers is a logical extension of their current roles in hematotherapy. In this paper, we describe the operation of one such centralized processing facility.

Locally applied hypothermia and microvascular muscle flap transfers.

The success rate for wound closure of grade III open tibia-fibula fractures with free muscle flaps is approximately 90%. Complications and loss of free flaps are due mainly to anastomotic problems, local anatomical considerations, or recipient vessel injuries, which prolong the ischemic time of the transferred free muscle tissue. We present the techniques used at the Shock Trauma Center of The Maryland Institute for Emergency Medical Services Systems (MIEMSS), which has allowed us a 100% success rate for the last 80 free muscle transfers performed. This surgical technique involves the use of locally applied hypothermia to decrease muscle metabolism and no-reflow phenomena. Representative cases are illustrated, which could have been failures because of increased ischemic time.

Skin banking: a simple method for cryopreservation of split-thickness skin and cultured human epidermal keratinocytes.

A simple unit has been developed for the simultaneous passive cooling of small and large amounts of allograft or autograft split-thickness skin, as well as cultured human epidermis. An expanded polystyrene box of variable size, aluminum plates, and cellulose tissue are fused. This unit is cooled in a -70 degrees C constant-temperature mechanical refrigerator. Maximal cooling rates of -1.3 degrees C min-1 are obtained in a box with a constant wall thickness of cellulose tissue. The cooling rate can be varied by altering the number of cellulose layers. Exothermic temperature plateaus associated with skin cooled in this unit last for less than 0.3 minutes. The viability of the cryopreserved skin was determined by using up to four methods simultaneously: a dye-exclusion test with trypan blue; glucose consumption; production of lactate; and carbon-13 nuclear magnetic resonance spectroscopy. Using a cryoprotectant medium with 15% (vol/vol) glycerol for split-thickness skin 0.25 mm thick, a storage time of up to 509 days at -70 degrees C was observed, with only a small decrease in viability (trypan blue, 62.5%; glucose consumption, 71 to 90% compared with freshly harvested skin). Storage in liquid nitrogen did not significantly improve results (p greater than 0.05).