Diagnostic accuracy of percutaneous biopsy in retroperitoneal sarcoma.

BACKGROUND: Percutaneous biopsy is recommended before surgery for suspected retroperitoneal sarcoma (RPS) to confirm the histological diagnosis and guide surgical strategy. The present study aimed to establish the diagnostic accuracy of percutaneous core biopsy with respect to histological diagnosis and tumour grade. METHODS: Data on patients with suspected RPS who underwent percutaneous biopsy followed by surgical resection between 2005 and 2016 at one of two tertiary European sarcoma units were reviewed. Histological tumour type and tumour grade on biopsy were correlated with postoperative histology to evaluate diagnostic accuracy. RESULTS: A total of 239 patients underwent percutaneous core biopsy followed by surgical resection in Milan (163, 68·2 per cent) or Birmingham (76, 31·8 per cent). Diagnostic accuracy varied with histological diagnosis (P < 0·001), but demonstrated overall concordance with final pathology following resection in 67·2 per cent of biopsies (κ = 0·606). The majority of discrepancies occurred in dedifferentiated liposarcoma (DDLPS), owing to under-recognition of dedifferentiation in this group. Concordance between pathology on biopsy and resection improved to 81·1 per cent when DDLPS and well differentiated liposarcoma were grouped together as liposarcoma. Grade on biopsy was concordant with grade on resection specimen in 60·4 per cent of tumours (κ = 0·640). Diagnosis of high-grade tumours on biopsy had a high specificity (98 per cent), and moderate positive predictive value (85 per cent) and negative predictive value (78 per cent). CONCLUSION: A diagnosis of DDLPS or leiomyosarcoma on percutaneous biopsy is highly reliable. High-grade sarcomas can be identified with high specificity, which opens the door to a study on neoadjuvant therapy in these patients.

Structural and functional 3D mapping of skin tumours with non-invasive multispectral optoacoustic tomography.

BACKGROUND: Recent advances in technology have enabled the development of various non-invasive skin imaging tools to aid real-time diagnosis of both benign and malignant skin tumours, minimizing the need for invasive skin biopsy. Multispectral optoacoustic tomography (MSOT) is a recently developed non-invasive imaging tool, which offers the unique capacity for high resolution three dimensional (3D) optical mapping of tissue by further delivering highly specific optical contrast from a depth of several millimetres to centimetres in living tissues. MSOT enables volumetric, spectroscopic differentiation of tissue, both in vivo and in real time, with and without the application of biomarker-specific probes, and is further able of providing spatial maps of skin chromophores, as well as underlying blood vasculature. METHODS: Three patients with suspicious skin tumours consented to have their lesions imaged with MSOT prior to excision. The histological findings and measurements were compared. RESULTS: We demonstrated the first in vivo clinical use of MSOT for 3D reconstruction of skin tumours in three patients with good histological correlation. CONCLUSION: Our findings confirm the potential benefit of the new imaging method in guiding surgical intervention to achieve a more precise excision with better clearance and lower relapse rates. It can also potentially help to shorten the duration of Mohs' micrographic surgery. Further large-scale studies are necessary to ensure correlation between MSOT and histology.

Measurement of proton tunneling in short hydrogen bonds in single crystals of 3,5 pyridinedicarboxylic acid using nuclear magnetic resonance spectroscopy.

In this Letter, we present NMR spin-lattice and relaxometry data for proton transfer in one of the shortest known N-H⋯O hydrogen bonds in a single crystal of 3,5 pyridinedicarboxylic acid (35PDCA). It is widely believed that proton transfer by quantum tunneling does not occur in short hydrogen bonds since the ground state energy level lies above the potential barrier, yet these data show a temperature independent, proton tunneling rate below 77 K and a clear deviation from classical dynamics below 91 K. This study therefore suggests that proton tunneling occurs in all hydrogen bonds at low temperature and the crossover temperature to classical hopping must be determined when evaluating whether proton tunneling persists at higher temperature, for example in enzyme catalysis under physiological conditions.

An optoacoustic imaging feature set to characterise blood vessels surrounding benign and malignant breast lesions.

Combining optoacoustic (OA) imaging with ultrasound (US) enables visualisation of functional blood vasculature in breast lesions by OA to be overlaid with the morphological information of US. Here, we develop a simple OA feature set to differentiate benign and malignant breast lesions. 94 female patients with benign, indeterminate or suspicious lesions were recruited and underwent OA-US. An OA-US imaging feature set was developed using images from the first 38 patients, which contained 14 malignant and 8 benign solid lesions. Two independent radiologists blindly scored the OA-US images of a further 56 patients, which included 31 malignant and 13 benign solid lesions, with a sensitivity of 96.8% and specificity of 84.6%. Our findings indicate that OA-US can reveal vascular patterns of breast lesions that indicate malignancy using a simple feature set based on single wavelength OA data, which is therefore amenable to application in low resource settings for breast cancer management.

The impact of postoperative radiological surveillance intensity on disease free and overall survival from primary retroperitoneal, abdominal and pelvic soft-tissue sarcoma.

AIM: This observational study aimed to evaluate the impact of intensity of radiological surveillance on survival following resection of retroperitoneal sarcoma. METHOD: Retrospective cohort study of patients undergoing primary resection of soft tissue sarcoma arising in the retroperitoneum, abdomen or pelvis at a single, high-volume sarcoma centre. Intensity of follow-up regimes up to 5 postoperative years were categorized as 'European Society for Medical Oncology (ESMO) compliant' (intense), or 'non-ESMO compliant' (less-intense). The primary outcome measure was overall survival (OS). The secondary outcome measures were disease-free survival (DFS) and reoperation rate. Analyses were stratified by high (grade 2 or 3) or low (grade 1) tumour grade. RESULTS: Of 168 patients, 67.1% had high-grade and 32.9% had low-grade disease. Overall, 40.0% of patients had ESMO-compliant radiological follow-up (high-grade:25.7%, low-grade:66.7%). 41.7% of patients died and 48.2% suffered local or distant recurrence by cessation of follow up. Upon univariable analysis for high-grade tumours, ESMO compliance reduced DFS (p = 0.066) but had no impact on OS. There was no significant difference in the reoperation rate in patients with ESMO-compliant and non-compliant follow-up (p = 0.097). In low-grade tumours, ESMO compliance significantly reduced DFS (p < 0.001), but without effecting OS. In risk-adjusted models for high-grade tumours, ESMO compliant follow-up was associated with reduced OS (HR:3.47, 1.40-8.61, p = 0.007) and no difference in DFS. In low-grade tumours, there was no association between overall ESMO compliance and OS or DFS. CONCLUSION: This study did not find a benefit for high-intensity radiological surveillance and overall survival in patients undergoing primary resection for high or low-grade retroperitoneal sarcoma.

Factors influencing selection for a day-case or 23-h stay procedure in transanal endoscopic microsurgery.

BACKGROUND: Transanal endoscopic microsurgery (TEMS) is an alternative to radical resection of the rectum for benign lesions and early rectal cancer. This study aimed to identify whether day-case TEMS is safe and which factors dictate patient suitability and length of stay (LOS). METHODS: Details of patients undergoing TEMS resection were retrieved from a tertiary referral prospective database. RESULTS: Of 96 patients, 46 (48 per cent) were day cases, 24 (25 per cent) had a 23-h stay and 26 (27 per cent) were inpatients. The frequency of day-case surgery increased significantly over the study interval (P = 0.050). Distance of the lesion from the anorectal junction, malignant potential and travel distance had no bearing on LOS. Older age (P = 0.004) and duration of surgery (P = 0.002) correlated significantly with increased LOS. Lesions covering one quadrant involved a significantly shorter stay than those covering two or more quadrants (P = 0.002). Maximum diameter (mean 5.7 cm) was strongly related to LOS (P = 0.009). Day-case and 23-h stay patients had a significantly higher proportion of lower-risk lesions (P = 0.001). CONCLUSION: High-volume day-case TEMS appears safe, even when long travel distances are involved. With advances in practice and procedural safety, traditional risk factors may not be as important as currently thought.

Gastrointestinal manifestations of extramedullary plasmacytoma: a narrative review and illustrative case reports.

Introduction Solitary extramedullary plasmacytoma are rare, solid-mass tumours which appear immunophenotypically similar to multiple myeloma. The diagnosis and management of gastrointestinal plasmacytoma is complex and requires multidisciplinary input. This study presents a narrative review of intra-abdominal extramedullary plasmacytoma, illustrated with two case studies. Methods The PubMed database was searched without date restrictions for reports of intra-abdominal extramedullary plasmacytoma to synthesise a narrative review. Electronic records were reviewed at a high-volume, quaternary soft-tissue sarcoma centre to identify patients with histopathologically confirmed extramedullary plasmacytoma affecting the gastrointestinal tract. Results Gastrointestinal extramedullary plasmacytomas can present with mass effect or organ-specific dysfunction. Techniques for tissue diagnosis of extramedullary plasmacytoma vary dependent on location, with a formal diagnosis often being made from a resected specimen. Management can include surgery, radiotherapy, systemic chemotherapy or a combination. No high-quality evidence base exists to guide treatment. Two case studies of operated gastrointestinal extramedullary plasmacytoma are presented at different phases of disease progression, with a resultant impact on survival. Conclusion Intra-abdominal extramedullary plasmacytoma is a rare and heterogeneous condition that lacks consensus guidelines for diagnosis and management. Collaboration between international specialist centres will create better quality evidence for treatment of this cohort.

An Update on Non-extremity Soft Tissue Sarcomas.

The management of soft tissue sarcoma is challenging and varied. Centralisation of management in high volume specialist centres has revolutionised outcomes. Surgery remains the mainstay of treatment and is currently the only potentially curative therapy. Retroperitoneal soft tissue sarcoma presents a particular challenge to the surgical oncologist and the concept of extended resection to include surrounding expendable organs taken en bloc with the tumour has now largely been adopted. The use of neoadjuvant and adjuvant therapies for retroperitoneal soft tissue sarcoma is still to be established, although they are employed on a case-specific basis. Guidance on the management of retroperitoneal recurrences and distant metastatic disease is now recognised. The approach to soft tissue sarcoma of the head and neck, trunk and abdominal wall remains largely extrapolated from experience of the management of extremity soft tissue sarcoma. Secondary angiosarcoma of the breast is becoming increasingly more common and presents a particular therapeutic challenge. Continued international collaboration is essential to ensure evolution of the optimal management of this rare group of cancers.

Role of palliative resection of the primary tumour in advanced pancreatic and small intestinal neuroendocrine tumours: A systematic review and meta-analysis.

PURPOSE: This study aimed to evaluate the impact on overall survival following palliative surgery to remove the primary lesion in unresectable metastatic small intestinal (SI-NET) and pancreatic neuroendocrine tumours (P-NET). METHODS: A systematic review of the literature and meta-analysis was performed. MEDLINE and Embase databases were searched to identify articles comparing patients undergoing palliative primary tumour resection without metastatectomy vs. no resection. Relevant articles were identified in accordance with PRISMA guidelines. The primary outcome was overall survival. Included studies were evaluated for heterogeneity and publication bias. RESULTS: 13 studies met the inclusion criteria, of which 6 presented data suitable for meta-analysis. No randomised controlled trials were identified. Analysis of pooled multivariate hazard ratios demonstrated significantly longer overall survival in patients undergoing resection of both P-NETs (HR 0.43; 95% CI: 0.34-0.57, p < 0.001) and SI-NETs (HR 0.47; 95% CI: 0.35-0.55, p = 0.007). The increase in median survival in patients treated surgically relative to non-surgically ranged from 14 to 46 months in P-NET, and 22-112 months in SI-NET. The number needed to treat in order that one additional patient was alive at five years, ranged from 3.0 to 4.2, and 1.7 to 7.7 respectively. CONCLUSIONS: Meta-analysis demonstrates that palliative resection of primary SI-NETs and P-NETs in the setting of unresectable metastatic disease can increase survival. Although these results should be interpreted with caution due to potential selection and publication bias, the data supports consideration of surgery, particularly in patients with low tumour burdens and good functional status.

Vibrational Raman optical activity of alpha-lactalbumin: comparison with lysozyme, and evidence for native tertiary folds in molten globule states.

Proteins in aqueous solution are now accessible to Raman optical activity (ROA) measurements, which provide an incisive new probe of secondary and tertiary structure illustrated here by a study of bovine alpha-lactalbumin. The room-temperature ROA spectrum of native bovine alpha-lactalbumin is similar to that of native hen egg-white lysozyme except for features attributable to differences in the loop regions: in particular, a positive ROA band at approximately 1338 cm-1 assigned to conformationally homogeneous loop structure, possibly with local order corresponding to 3(10)-helix, has more than double the intensity in alpha-lactalbumin compared with lysozyme. This is consistent with the two proteins having similar secondary structure but different local details in the tertiary fold. ROA measurements on alpha-lactalbumin at pH 2.0 over a range of temperatures have provided a new perspective on the molten globule state. Thus at 35 degrees C ROA reveals the presence of some secondary structure but an almost complete loss of the tertiary loop structure; whereas at 2 degrees C the ROA spectrum is almost identical with that of the native protein, which is strong evidence that virtually all of the secondary structure and the tertiary backbone fold persist, albeit within a looser framework associated with increased solvent exposure and change of environment of many of the side-chains as evidenced by an increase in noise and bandwidth of some of the ROA signals together with aromatic fluorescence and near-UV circular dichroism signals characteristic of the molten globule state. Our sample of acid alpha-lactalbumin at 2 degrees C therefore appears to be an archetypal example of Ptitsyn's "native-like" molten globule, having a fixed native-like tertiary fold but with loss of tight packing of the side-chains; whereas at 35 degrees C it is a "disordered" molten globule. At 20 degrees C the acid molten globule appears to retain highly native-like secondary structure but with most of the tertiary fold already lost. A calcium-free sample of alpha-lactalbumin at neutral pH displayed a broad cooperative transition between native and molten globule states at approximately 15 degrees C, with the latter state showing similar but somewhat degraded tertiary loop ROA signatures to the native protein. In both the acid and apo molten globule states the ROA signatures of the secondary structure and the tertiary loops showed a gradual change with temperature.

Use of mathematical derivatives (time-domain differentiation) on chromatographic data to enhance the detection and quantification of an unknown 'rider' peak.

Two samples of an anticancer prodrug, AQ4N, were submitted for HPLC assay and showed an unidentified impurity that eluted as a 'rider' on the tail of the main peak. Mathematical derivatization of the chromatograms offered several advantages over conventional skimmed integration. A combination of the second derivative amplitude and simple linear regression gave a novel method for estimating the true peak area of the impurity peak. All the calculation steps were carried out using a widely available spreadsheet program.

Development of a lyophilised RH1 formulation: a novel DT diaphorase activated alkylating agent.

RH1 is a novel aziridinylbenzoquinone alkylating agent, which is activated in tumour cells by DT diaphorase. In common with previous aziridinylbenzoquinones, RH1 exhibits limited aqueous stability and solubility. The aim of this study was to examine the pharmaceutical properties of RH1 with a view to preparing a suitable formulation for clinical trial. Stability in a neutral phosphate-buffered solution was poor with a degradation half-life of 50 h at 55 degrees C, indicating that lyophilisation was preferable. The reaction kinetics indicated a similarity with previous studies for base-catalysed degradation of aziridinylbenzoquinones. Intrinsic aqueous solubility at 0.5 mg mL(-1) may be increased in solvent systems or by the use of polymers such as polyvinylpyrrolidone (PVP) or complexing agents like hydroxypropyl-beta-cyclodextrin (HPBCD). In the latter case this increased solubility by an order of magnitude to around 5 mg mL(-1). Four potential formulations based on lyophilisation of RH1 (1 mg mL(-1)) from buffered solution (pH 7, 0.01 M NaH2PO4) containing either 50 mg mL(-1) mannitol, 40 mg mL(-1) dextran, 20 mg mL(-1) PVP or 50 mg mL(-1) HPBCD were prepared and examined for stability characteristics. All formulations exhibited a temperature-dependent degradation. The mannitol and dextran formulations had limited stability and degraded rapidly at all temperatures. The PVP and HPBCD formulations degraded at elevated temperatures but remained stable for up to twelve months at 4 degrees C. Examination of the degradation kinetics in the latter systems demonstrated similarity to the solution degradation mechanism, while in the former alternative degradation pathways appeared to be occurring. The chemical stability of RH1 in lyophilised formulations is dependent upon the excipient employed and storage temperature. Either the PVP or HPBCD formulation would be suitable clinical trial formulations of RH1. The results indicate that the choice of lyophilisation excipient for aziridinylbenzoquinones cannot be based on previous literature studies of related agents.

Vibrational Raman optical activity of glycoproteins.

This paper reports the first vibrational Raman optical activity (ROA) spectrum of a glycoprotein. The sample, orosomucoid (alpha 1-acid glycoprotein), shows ROA bands characteristic of a high beta-sheet content together with new bands which could be specific for the carbohydrate and its association with the protein. Our results suggest that ROA spectra of intact glycoproteins may contain information about both protein and carbohydrate conformation and the mutual influence on each other's stability and conformation.

Equine rhinopneumonitis virus (herpesvirus type 1): attenuation in stable monkey cell line.

An isolate of virulent equine herpesvirus (EHV) type 1 was adapted to Vero stable cell line by 13 serial passages at 37 C and 50 serial passages at 26 C. Characteristics of the attenuated EHV-1 were found to be avirulent, but immunogenic in horses if injected intramuscularly. The attenuated virus was regularly isolated from peripheral leukocytes in inoculated horses, but was not recovered from nasal turbinate tissues. A mild leukopenia was noticed. The attenuated virus produced characteristic large syncytia on primary isolation in rabbit kidney (RK13) or Vero cells at 37 C in contrast to cell rounding observed with virulent EHV-1. The syncytial marker was stable through 20 serial passages in Vero cells at 37 C. New application of double immunodiffusion test for distinguishing between EHV-1 and EHV-2 also is described.

A comparative study of the iron status of patients with oesophageal adenocarcinoma to determine suitability for a clinical trial of iron chelation therapy.

INTRODUCTION: The incidence of oesophageal adenocarcinoma (OAC) is rising dramatically and overall survival remains extremely poor. Iron has been shown to potentiate tumourigenesis in OAC, and iron chelation therapy demonstrates promise in vivo as an adjunct to neoadjuvant and palliative chemotherapy. OAC, however, has traditionally been associated with iron deficiency anaemia. The aim of this study was therefore to formally quantify the iron status of OAC patients in order to guide the design of future clinical trials involving iron chelation therapy. METHODS: Demographic and cancer specific data were collected prospectively from all patients presenting with OAC and gastric adenocarcinoma (GAC). Patients had haemoglobin, serum iron, serum ferritin and serum transferrin receptor (sTfR) levels measured to assess systemic iron status. In addition, the sTfR/log ferritin (sTfR-F) index was calculated. RESULTS: Average haemoglobin, serum iron, serum ferritin, sTfR and sTfR-F index values for all patients presenting with OAC were within normal sex specific reference ranges. No statistical difference in iron status was observed between OAC patients presenting with resectable and advanced OAC. Patients with OAC are relatively iron replete compared with those presenting with GAC. Iron parameters were not significantly altered by standard neoadjuvant chemotherapy. CONCLUSIONS: Patients presenting with resectable or advanced OAC could be considered as candidates for a clinical trial of iron chelation therapy as an addition to standard neoadjuvant or palliative treatments.

The implementation and effectiveness of an enhanced recovery programme after oesophago-gastrectomy: a prospective cohort study.

BACKGROUND: Oesophageal resection is notoriously complicated and produces a cohort of patients prone to postoperative complications. Maintaining quality care demands a systematic approach to patient management yet postoperative recovery after oesophagectomy is often needlessly inefficient, heterogeneous and governed by the idiosyncrasies of the operating surgeon. Enhanced recovery after surgery (ERAS) programmes are now well established in colorectal surgery and here we describe the implementation and effectiveness of an ERAS programme for the postoperative management of Ivor Lewis oesophago-gastrectomy (ILOG). METHODS: An ERAS programme was devised and implemented with the support of a dedicated in-hospital task-force. Three consultant surgeons allocated consecutive patients to the programme (ERAS) and outcomes were compared to consecutive patients not on the ERAS programme (non-ERAS) and a pre-ERAS cohort (pre-ERAS). Principal outcome measures were total length of stay (TLOS), Accordion postoperative complication grade and 30-day readmission rate. RESULTS: 75 patients were enrolled on the ERAS programme, 41 continued as a non-ERAS cohort and 80 consecutive pre-ERAS patients were identified. A significant improvement in median TLOS was observed in the ERAS group (10 days r.7-58) compared to pre-ERAS (13 days r. 8-57) (p = <0.001) and non-ERAS patients (13 days r.8-42) (p = <0.001). No significant difference in Accordion scores for postoperative complications or 30-day readmission rates were observed. DISCUSSION: The introduction of an ERAS programme after ILOG can significantly reduce TLOS without jeopardising patient safety or clinical outcomes. The successful introduction of an ERAS programme requires full motivation and support from all team members including the patient.

Deferasirox (ICL670A) effectively inhibits oesophageal cancer growth in vitro and in vivo.

BACKGROUND AND PURPOSE: Growing evidence implicates iron in the aetiology of gastrointestinal cancer. Furthermore, studies demonstrate that iron chelators possess potent anti-tumour activity, although whether iron chelators show activity against oesophageal cancer is not known. EXPERIMENTAL APPROACH: The effect of the iron chelators, deferoxamine (DFO) and deferasirox, on cellular iron metabolism, viability and proliferation was assessed in two oesophageal adenocarcinoma cell lines, OE33 and OE19, and the squamous oesophageal cell line, OE21. A murine xenograft model was employed to assess the effect of deferasirox on oesophageal tumour burden. The ability of chelators to overcome chemoresistance and to enhance the efficacy of standard chemotherapeutic agents (cisplatin, fluorouracil and epirubicin) was also assessed. KEY RESULTS: Deferasirox and DFO effectively inhibited cellular iron acquisition and promoted intracellular iron mobilization. The resulting reduction in cellular iron levels was reflected by increased transferrin receptor 1 expression and reduced cellular viability and proliferation. Treating oesophageal tumour cell lines with an iron chelator in addition to a standard chemotherapeutic agent resulted in a reduction in cellular viability and proliferation compared with the chemotherapeutic agent alone. Both DFO and deferasirox were able to overcome cisplatin resistance. Furthermore, in human xenograft models, deferasirox was able to significantly suppress tumour growth, which was associated with decreased tumour iron levels. CONCLUSIONS AND IMPLICATIONS: The clinically established iron chelators, DFO and deferasirox, effectively deplete iron from oesophageal tumour cells, resulting in growth suppression. These data provide a platform for assessing the utility of these chelators in the treatment of oesophageal cancer patients.

Pharmaceutical development of the novel arsenical based cancer therapeutic GSAO for Phase I clinical trial.

The novel organoarsenical GSAO, 4-(N-(S-glutathionylacetyl)amino) phenylarsonous acid, has potential anti-angiogenic capability with application in cancer where tumour metastasis relies on neo-vascularisation. As GSAO arsenic is trivalent, the arsenoxide moiety reacts with appropriately spaced cysteine residues on adenine nucleotide translocase (ANT) mitochondrial membrane protein. Molecular oxidation of the arsenic to the pentavalent structure, as in the degradant GSAA (4-(N-(S-glutathionylacetyl)amino) phenylarsonic acid), prevents sulphydryl interaction and risks abolition of activity. We report here on formulation studies aiming to produce a parenteral product with the primary objective of restricting GSAA transformation from GSAO to protect maximal potency of the molecule. Successful anti-oxidant strategy primarily came from pH control. The presence of glycine was proposed to form a stabilising five-membered oxazarsolidinone ring with arsenoxide and this was investigated using potentiometric assays. We report on these tritration studies identifying a pK(a) of 8.2 associated with an As-OH, but not confirming ring presence. An original clinical trial pharmaceutical was successfully realised by lyophilisation of 50 mg/mL GSAO in 100 mM glycine solution, pH 7 to obtain a 48-month shelf life for the freeze-dried vials. The Phase I clinical study is ongoing in patients with solid tumours refractory to standard therapy.

Stabilization of bacteriophage during freeze drying.

With preliminary clinical trials completed for the treatment of antibiotic resistant infections using bacteriophages, there is a need to develop pharmaceutically acceptable formulations. Lyophilization is an established technique for the storage of bacteriophage, but there is little consensus regarding drying cycles, additives and moisture content specific to phage. Here, the addition of sucrose or poly(ethylene glycol) 6000 yielded stable freeze-dried cakes only from high concentrations (0.5 M and 5%, respectively), with addition of bacteriophage otherwise causing collapse. Gelatin, which is added to storage media (a solution of salts), played no role in maintaining bacteriophage stability following lyophilization. A secondary drying cycle was most important for maintaining bacteriophage activity. The addition of high concentrations of PEG 6000 or sucrose generally caused a more rapid fall in bacteriophage stability, over the first 7-14 d, but thereafter residual activities for all phage formulations converged. There was no distinct change in the glass transition temperatures (T(g)) measured for the formulations containing the same additive. Imaging of cakes containing fluorescently labeled bacteriophage did not show gross aggregation or phase separation of bacteriophage during lyophilization. However, the moisture content of the cake did correlate with lytic activity, irrespective of the formulation, with a 4-6% moisture content proving optimal. We propose that residual moisture is followed during lyophilization of bacteriophage from minimal concentrations of bulking agent.