Optimizing genomic prediction for Australian Red dairy cattle.

The reliability of genomic prediction is influenced by several factors, including the size of the reference population, which makes genomic prediction for breeds with a relatively small population size challenging, such as Australian Red dairy cattle. Including other breeds in the reference population may help to increase the size of the reference population, but the reliability of genomic prediction is also influenced by the relatedness between the reference and validation population. Our objective was to optimize the reference population for genomic prediction of Australian Red dairy cattle. A reference population comprising up to 3,248 Holstein bulls, 48,386 Holstein cows, 807 Jersey bulls, 8,734 Jersey cows, and 3,041 Australian Red cows and a validation population with between 208 and 224 Australian Red Bulls were used, with records for milk, fat, and protein yield, somatic cell count, fertility, and survival. Three different analyses were implemented: single-trait genomic best linear unbiased predictor (GBLUP), multi-trait GBLUP, and single-trait Bayes R, using 2 different medium-density SNP panels: the standard 50K chip and a custom array of variants that were expected to be enriched for causative mutations. Various reference populations were constructed containing the Australian Red cows and all Holstein and Jersey bulls and cows, all Holstein and Jersey bulls, all Holstein bulls and cows, all Holstein bulls, and a subset of the Holstein individuals varying the relatedness between Holsteins and Australian Reds and the number of Holsteins. Varying the relatedness between reference and validation populations only led to small changes in reliability. Whereas adding a limited number of closely related Holsteins increased reliabilities compared with within-breed prediction, increasing the number of Holsteins decreased the reliability. The multi-trait GBLUP, which considered the same trait in different breeds as correlated traits, yielded higher reliabilities than the single-trait GBLUP. Bayes R yielded lower reliabilities than multi-trait GBLUP and outperformed single-trait GBLUP for larger reference populations. Our results show that increasing the size of a multi-breed reference population may result in a reference population dominated by one breed and reduce the reliability to predict in other breeds.

Social impairments in autism spectrum disorder are related to maternal immune history profile.

Maternal immune activation has been highlighted as a factor that might increase the risk and severity of autism spectrum disorder (ASD) in children. Preclinical animal evidence shows that immune activation in mothers during pregnancy causes ASD-like behavioural traits in offspring. To this point, there has been no investigation of whether immune system activation in human mothers during pregnancy is associated with more severe symptoms in children with ASD. In this study, data from an existing ASD cohort (N=220) were analysed to investigate whether immune conditions in the mother were associated with greater severity of autism-related symptoms. Results showed that children whose mothers reported a history of immune activation (allergies and asthma) had significantly higher scores on the Social Responsiveness Scale (SRS; P=0.016), suggesting more severe social impairment symptoms in these children. This increasing severity of social impairment symptoms was further shown on the SRS cognition (P=0.007) and mannerisms (P=0.002) subscales. While immune history was associated with an increase in the severity of social impairment symptoms, history of autoimmune conditions in the mother did not have any effect in this cohort. To the best of our knowledge, this study is the first to show an association between immune activation history in the mother and increased ASD symptom severity in children with ASD. These findings support the idea of an immune system-mediated subtype in ASD, where the immune history of the mother may be an important factor.

Preexisting mild sleep disturbance as a vulnerability factor for inflammation-induced depressed mood: a human experimental study.

Sleep disturbance and depression are common, particularly in females, and sleep disturbance is a well-known risk factor for depression. Systemic inflammation has been suggested as a potential mechanism of this association. This study examined whether preexisting sleep disturbance acted as a vulnerability factor for depressed mood induced by an inflammatory challenge in healthy females vs males. In a randomized double-blind placebo-controlled design, volunteers aged 18-50 (N = 111; 67 females) were assigned to placebo or low-dose endotoxin. Before substance administration, sleep disturbance was assessed using the Pittsburgh Sleep Quality Index and dichotomized using median split (⩾ 3 vs < 3). Self-reported depressed mood (profile of mood states) and circulating proinflammatory cytokines (interleukin-6, tumor necrosis factor-α) were repeatedly assessed over 6 h. Among females, moderation of depressed mood by sleep disturbance was significant even after adjustment for covariates (X(2) = 12.73, df = 6, P < 0.05). There was a robust time-by-condition interaction in females with sleep disturbance (X(2) = 26.22, df = 6, P < 0.001), but not in females without sleep disturbance (X(2) = 8.65, df = 6, P = 0.19). Although cytokines increased equally in all females, the correlations between cytokines and depressed mood were significantly stronger in females with sleep disturbance. Among males, no moderating effect of sleep disturbance was observed. Inflammation-induced depressed mood was considerably more severe among females reporting mild sleep disturbance compared with those reporting no sleep disturbance, suggesting that even mild sleep disturbance may increase vulnerability for inflammation-induced depression in females. Furthermore, sleep disturbance appears to increase the vulnerability to depression by augmenting affective sensitivity to cytokines rather than by enhancing cytokine responses to inflammatory challenge in females.

Understanding STAT3 signaling in cardiac ischemia.

Cardiovascular disease is the leading cause of death worldwide. It remains one of the greatest challenges to global health and will continue to dominate mortality trends in the future. Acute myocardial infarction results in 7.4 million deaths globally per annum. Current management strategies are centered on restoration of coronary blood flow via percutaneous coronary intervention, coronary artery bypass grafting and administration of anti-platelet agents. Such myocardial reperfusion accounts for 40-50 % of the final infarct size in most cases. Signaling transducer and activator of transcription 3 (STAT3) has been shown to have cardioprotective effects via canonical and non-canonical activation and modulation of mitochondrial and transcriptional responses. A significant body of in vitro and in vivo evidence suggests that activation of the STAT3 signal transduction pathway results in a cardio protective response to ischemia and attempts have been made to modulate this with therapeutic effect. Not only is STAT3 important for cardiomyocyte function, but it also modulates the cardiac microenvironment and communicates with cardiac fibroblasts. To this end, we here review the current evidence supporting the manipulation of STAT3 for therapeutic benefit in cardiac ischemia and identify areas for future research.

The association between pro-inflammatory cytokines, regional cerebral metabolism, and cognitive complaints following adjuvant chemotherapy for breast cancer.

To examine relationships following adjuvant chemotherapy between circulating pro-inflammatory cytokines, regional cerebral metabolism, and cognitive complaints in early stage breast cancer patients. 33 breast cancer patients who had completed initial treatment (surgery, ± radiation, 23 chemotherapy, 10 no chemotherapy) obtained resting (18)F-FDG PET/CT brain imaging at baseline and 1 year later. Pro-inflammatory cytokine markers (IL-1ra, sTNF-RII, CRP, and IL-6) and cognitive complaints were also assessed at both time points. At baseline, consistent correlations were seen between the left medial frontal and right inferior lateral anterior temporal cortices and inflammatory markers within the chemotherapy group, and not in the no chemotherapy group. After 1 year, correlations persisted in the medial frontal cortex and the temporal cortex, the latter shifting superiorly. Both of these regional correlations demonstrated the highest levels of significance when looking across the 1 year time frame (IL-1ra: peak voxel p < 0.0005; cluster size p < 0.0005, p = 0.001 after correction (medial prefrontal), p < 0.0005; cluster size p = 0.001, p = 0.029 corr. (anterior temporal), sTNF-RII: p < 0.0005; cluster size p = 0.001, p = 0.040 corr. (medial prefrontal)). Positive correlations were also seen within the chemotherapy group between baseline memory complaints and the medial frontal (p < 0.0005; cluster size p < 0.0005, p < 0.0005 corr.) and anterior temporal (p < 0.0005; cluster size p < 0.0005, p = 0.002 corr.) cortices at baseline and 1 year later. Metabolism in the medial prefrontal cortex and anterior temporal cortex was found to correlate with both memory complaints and cytokine marker levels in chemotherapy patients.

Does tumor necrosis factor-alpha (TNF-α) play a role in post-chemotherapy cerebral dysfunction?

Post-chemotherapy treated cancer patients frequently report cognitive difficulties. The biology of this phenomenon is poorly understood, with uncertainty about possible direct toxic effects on the brain, secondary effects from systemic inflammation, host factors/genetic predisposition to cognitive complaints, or hormonal changes influencing cognitive function. To elucidate possible mechanisms associated with post-treatment cognitive dysfunction among breast cancer survivors, in 2007 we established a prospective, longitudinal, observational cohort study of early stage breast cancer patients, recruited at the end of initial treatments (primary treatment exposure included surgery, ± radiation, ± chemotherapy), and prior to the initiation of adjuvant endocrine therapy. We assessed cognitive complaints, neuropsychological (NP) test performance, markers of inflammation, and brain imaging at baseline, 6 months and 12 months after enrollment. In this analysis of data from the first 93 patients enrolled in the cohort study, we focus on the relationship of circulating levels of proinflammatory cytokines to cerebral functioning and chemotherapy exposure. Among the proinflammatory cytokines tested (IL-1 ra, sTNF-RII, CRP, and IL-6) at baseline, only sTNF-RII was increased among chemotherapy exposed patients, with a significant decline in the year after treatment (p=0.003). Higher baseline sTNF-RII in chemotherapy patients was significantly associated with increased memory complaints. In chemotherapy exposed patients, the longitudinal decline in sTNF-RII was significantly correlated with fewer memory complaints over 12 months (r=-0.34, p=0.04). Higher baseline sTNF-RII was also associated with relatively diminished brain metabolism in the inferior frontal cortex (r=-0.55, p=0.02), as well as relatively increased inferior frontal metabolism after 1 year, in chemotherapy-exposed subjects. These preliminary findings suggest that post-chemotherapy increases in TNF-α may be playing an important role in the manifestations of cognitive complaints in breast cancer survivors.

Tubulin-targeting agents in hybrid drugs.

The targeting of tubulin is an important mechanism for cancer chemotherapy. However, limitations such as resistance, toxicity and incomplete tumour elimination associated with individual anti-cancer drugs have led to a need for combination therapy in cancer. It is therefore relevant to ask whether two or more drugs might be combined in a single hybrid molecule to advantageous effect. This review provides an overview of the hybrid drugs thus far investigated, in which at least one component targets tubulin. The rationale behind this approach is that the hybrid drug may have activity enhanced above and beyond that of the equivalent drug combination, or have an otherwise improved clinical outcome. Particular emphasis is placed on the investigation of activity in multidrug-resistant cancer cell lines. Attention is drawn to the difficulties encountered when developing hybrid drugs, with respect to in vivo metabolism-tracking, increased molecular bulk, and optimisation of the drug dosage ratio. The actual and potential advantages and disadvantages of such hybrid drugs when compared to single drugs or drug combinations are discussed critically and promising directions for future research is highlighted.

Paracetamol availability in pharmacy and non-pharmacy outlets in Dublin, Ireland.

INTRODUCTION: In 2004, there were 11,092 presentations to Irish hospitals with deliberate self-harm, including 7,933 cases of drug overdose, of which 31% involved paracetamol. Limiting the availability of paracetamol reduces morbidity and mortality associated with paracetamol overdose. AIM: The present study aimed to determine the level of compliance with statutory regulations governing the sale of paracetamol in Ireland. METHODS: Researchers visited pharmacy (n = 20) and non-pharmacy outlets (newsagents, mini-markets and supermarkets) (n = 50) in Dublin city and attempted to purchase amounts of paracetamol that exceeded the statutory limits for a single transaction. RESULTS: Amounts of paracetamol in excess of statutory limits for a single transaction were purchased in 50.0% of pharmacies, 81.8% of newsagents/mini-markets and 20.0% of supermarkets. One year later, we again visited pharmacy (n = 20) and non-pharmacy outlets (n = 50) in Dublin city and purchased amounts of paracetamol in excess of statutory limits in 50.0% of pharmacies, 52.3% of newsagents/mini-markets and 10.0% of supermarkets. CONCLUSION: We recommend that (a) notwithstanding the improvement in compliance rates in newsagents/mini-markets, the sale of paracetamol in these outlets should be discontinued; (b) the sale of paracetamol in supermarkets should continue, although automated checkout tills should be appropriately re-programmed; and (c) there should be greater efforts to ensure compliance with statutory regulations in pharmacies.

Immunodetection of UCP1 in rat thymocytes.

Thymi were dissected from rats and connective tissue was removed. Mitochondria were purified from isolated thymocytes and immunoblot analysis was performed using an antibody specific for uncoupling protein 1, which detected a 32.5 kDa protein associated with mitochondria from the thymocytes. This implies that rat thymocytes contain uncoupling protein 1.

How complete and accurate is meningococcal disease notification?

Effective public health control of meningococcal disease (meningococcal meningitis and septicaemia) is dependent on complete, accurate and speedy notification. Using capture-recapture techniques this study assesses the completeness, accuracy and timeliness of meningococcal notification in a health authority. The completeness of meningococcal disease notification was 94.8% (95% confidence interval 93.2% to 96.2%); 91.2% of cases in 2001 were notified within 24 hours of diagnosis, but 28.0% of notifications in 2001 were false positives. Clinical staff need to be aware of the public health implications of a notification of meningococcal disease, and of failure of, or delay in notification. Incomplete or delayed notification not only leads to inaccurate data collection but also means that important public health measures may not be taken. A clinical diagnosis of meningococcal disease should be carefully considered between the clinician and the consultant in communicable disease control (CCDC). Otherwise, prophylaxis may be given unnecessarily, disease incidence inflated, and the benefits of control measures underestimated. Consultants in communicable disease control (CCDCs), in conjunction with clinical staff, should de-notify meningococcal disease if the diagnosis changes.

Selective detection of UCP 3 expression in skeletal muscle: effect of thyroid status and temperature acclimation.

A novel peptide antibody to UCP 3 is characterized which is sensitive and discriminatory for UCP 3 over UCP 2, UCP 1 and other mitochondrial transporters. The peptide antibody detects UCP 3 expression in E. coli, COS cells and yeast expression systems. The peptide antibody detects a single approximately 33 kDa protein band in mitochondria from isolated rat skeletal muscle, mouse and rat brown adipose tissue, and in whole muscle groups (soleus and extensor digitorum longus) from mice. No 33 kDa band is detectable in isolated mitochondria from liver, heart, brain, kidney and lungs of rats, or gastrocnemius mitochondria from UCP 3 knock-out mice. From our data, we conclude that the peptide antibody is detecting UCP 3 in skeletal muscle, skeletal muscle mitochondria and brown adipose tissue mitochondria. It is also noteworthy that the peptide antibody can detect human, mouse and rat forms of UCP 3. Using the UCP 3 peptide antibody, we confirm and quantify the increased (2.8-fold) UCP 3 expression observed in skeletal muscle mitochondria isolated from 48-h-starved rats. We show that UCP 3 expression is increased (1.6-fold) in skeletal muscle of rats acclimated over 8 weeks to 8 degrees C and that UCP 3 expression is decreased (1.4-fold) in rats acclimated to 30 degrees C. Furthermore, UCP 3 expression is increased (2.3-fold) in skeletal muscle from hyperthyroid rats compared to euthyroid controls. In addition, we show that UCP 3 expression is only coincident with the mitochondrial fraction of skeletal muscle homogenates and not peroxisomal, nuclear or cytosolic and microsomal fractions.

Viral interleukin 6 stimulates human peripheral blood B cells that are unresponsive to human interleukin 6.

Cellular responsiveness to human interleukin 6 (hIL6) requires the expression of two receptor molecules: IL6-specific receptor (CD126'IL6R') and a nonspecific signal-transducing molecule (CD130'gp130'). Regulation of responsiveness to hIL6 is generally controlled by CD126'IL6R' expression. A viral homologue of hIL6 (vIL6) is encoded by human herpesvirus-8 and has biologic activity similar to hIL6 on a number of cell lines. vIL6 differs from hIL6 in its receptor utilization, requiring only CD130'gp130'. Total human B cells isolated from peripheral blood, which are predominantly CD126'IL6R'-negative, as well as sorted CD126'IL6R'-negative B cells, could be stimulated by recombinant vIL6, but not by hIL6, as indicated by induction of IL6-like signaling (STAT3 phosphorylation). This suggests that the ability of vIL6 to stimulate B cells expressing little or no CD126'IL6R' allows it to act on a larger pool of target B cells, compared to human IL6.

Effect of moisture on the stability of a lyophilized humanized monoclonal antibody formulation.

PURPOSE: To determine the effect of moisture and the role of the glass transition temperature (Tg) on the stability of a high concentration, lyophilized, monoclonal antibody. METHODS: A humanized monoclonal antibody was lyophilized in a sucrose/histidine/polysorbate 20 formulation. Residual moistures were from 1 to 8%. Tg values were measured by modulated DSC. Vials were stored at temperatures from 5 to 50 degrees C for 6 or 12 months. Aggregation was monitored by size exclusion chromatography and Asp isomerization by hydrophobic interaction chromatography. Changes in secondary structure were monitored by Fourier transform infrared (FTIR). RESULTS: T. values varied from 80 degrees C at 1% moisture to 25 degrees C at 8% moisture, there was no cake collapse and were no differences in the secondary structure by FTIR. All formulations were stable at 5 degrees C. High moisture cakes had higher aggregation rates than drier samples if stored above their Tg values. Intermediate moisture vials were more stable to aggregation than dry vials. High moisture samples had increased rates of Asp isomerization at elevated temperatures both above and below their Tg values. Chemical and physical degradation pathways followed Arrhenius kinetics during storage in the glassy state. Only Asp isomerization followed the Arrhenius model above the Tg value. Both chemical and physical stability at T > or = Tg were fitted to Williams-Landel-Ferry (WLF) kinetics. The WLF constants were dependent on the nature of the degradation system and were not characteristic of the solid system. CONCLUSION: High moisture levels decreased chemical stability of the formulation regardless of whether the protein was in a glassy or rubbery state. In contrast, physical stability was not compromised, and may even be enhanced, by increasing residual moisture if storage is below the Tg value.

Skeletal muscle capillarity and angiogenic mRNA levels after exercise training in normoxia and chronic hypoxia.

Gene expression of vascular endothelial growth factor (VEGF), and to a lesser extent of transforming growth factor-beta(1) (TGF-beta(1)) and basic fibroblast growth factor (bFGF), has been found to increase in rat skeletal muscle after a single exercise bout. In addition, acute hypoxia augments the VEGF mRNA response to exercise, which suggests that, if VEGF is important in muscle angiogenesis, hypoxic training might produce greater capillary growth than normoxic training. Therefore, we examined the effects of exercise training (treadmill running at the same absolute intensity) in normoxia and hypoxia (inspired O(2) fraction = 0.12) on rat skeletal muscle capillarity and on resting and postexercise gene expression of VEGF, its major receptors (flt-1 and flk-1), TGF-beta(1), and bFGF. Normoxic training did not alter basal or exercise-induced VEGF mRNA levels but produced a modest twofold increase in bFGF mRNA (P < 0.05). Rats trained in hypoxia exhibited an attenuated VEGF mRNA response to exercise (1.8-fold compared 3.4-fold with normoxic training; P < 0.05), absent TGF-beta(1) and flt-1 mRNA responses to exercise, and an approximately threefold (P < 0.05) decrease in bFGF mRNA levels. flk-1 mRNA levels were not significantly altered by either normoxic or hypoxic training. An increase in skeletal muscle capillarity was observed only in hypoxically trained rats. These data show that, whereas training in hypoxia potentiates the adaptive angiogenic response of skeletal muscle to a given absolute intensity of exercise, this was not evident in the gene expression of VEGF or its receptors when assessed at the end of training.

Diaphragmatic angiogenic growth factor mRNA responses to increased ventilation caused by hypoxia and hypercapnia.

This study investigates the effect of increased ventilation on the expression of messenger ribonucleic acid (mRNA) levels of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and transforming growth factor-beta1 (TGF-beta1) in the diaphragm of intact, awake, spontaneously breathing rats, compared with responses in paralysed, mechanically-ventilated animals at similar blood gas and ventilatory levels. Four groups of intact, rats were studied in a body box, each group breathing one of four gases: room air, 12% oxygen (O2), 5% carbon dioxide (CO2), or 12% O2+5% CO2 for 1 h. Another 4 groups of paralysed, mechanically-ventilated animals were matched for arterial blood gas and ventilatory level. The results showed that VEGF mRNA abundance was increased three-fold and that of bFGF 1.5-fold when 12% O2+5% CO2 were breathed, but TGF-beta1 did not change. A significant linear relationship of VEGF and bFGF mRNA to minute ventilation was observed in awake animals (r=0.98, p<0.02 and r=0.87, p<0.03, respectively). The paralysed, mechanically-ventilated animals showed no mRNA increases for any probe. Systemic hypoxia had no additional effect on VEGF or bFGF levels in the diaphragm. It was concluded that messenger ribonucleic acid for vascular endothelial growth factor and basic fibroblast growth factor in the diaphragm rises significantly as a result of active ventilation and not due to blood gas/pH changes or to passive muscle shortening per se.

Chronic hypoxia attenuates resting and exercise-induced VEGF, flt-1, and flk-1 mRNA levels in skeletal muscle.

Vascular endothelial growth factor (VEGF) is a hypoxia-inducible angiogenic mitogen. However, chronic hypoxia is generally not found to increase mammalian skeletal muscle capillarity. We sought to determine the effect of chronic hypoxia (8 wk, inspired O2 fraction = 0.12) on skeletal muscle gene expression of VEGF, its receptors (flt-1 and flk-1), basic fibroblast growth factor, and transforming growth factor-beta1. Wistar rats were exposed to chronic hypoxia (n = 12) or room air (n = 12). After the exposure period, six animals from each group were subjected to a single 1-h treadmill exercise bout (18 m/min on a 10 degrees incline) in room air while the remaining six animals served as rest controls. Morphological analysis revealed that chronic hypoxia did not increase skeletal muscle capillarity. Northern blot analyses showed that chronic hypoxia decreased resting VEGF, flt-1, and flk-1 mRNA by 23, 68, and 42%, respectively (P < 0.05). The VEGF mRNA response to exercise was also decreased (4.1- and 2.7-fold increase in room air and chronic hypoxia, respectively, P < 0.05). In contrast, neither transforming growth factor-beta1 nor basic fibroblast growth factor mRNA was significantly altered by chronic hypoxia. In conclusion, prolonged exposure to hypoxia attenuated gene expression of VEGF and its receptors flt-1 and flk-1 in rat gastrocnemius muscle. These findings may provide an explanation for the lack of mammalian skeletal muscle angiogenesis that is observed after chronic hypoxia.

Cytokine gene expression occurs more rapidly in stimulated peripheral blood mononuclear cells from human immunodeficiency virus-infected persons.

Evaluation of cytokine gene expression following in vitro stimulation is one means of examining the dysregulation of the immune system in human immunodeficiency virus (HIV) infection. We have assessed differences in the immune status of non-HIV-infected (HIV-) and HIV-infected (HIV+) individuals by evaluating the kinetics of the expression of cytokine genes. We compared detailed time courses of cytokine mRNA expression in HIV- and HIV+ peripheral blood mononuclear cells (PBMC) and found that there is a significant shift (P<0.01) for all cytokines examined (interleukin 2 [IL-2], IL-6, IL-10, gamma interferon, and tumor necrosis factor alpha [TNF-alpha]) to an earlier time of mean peak mRNA expression by HIV+ PBMC (between 4 and 8 h) compared to HIV- PBMC (8 h) in response to either phytohemagglutinin (PHA) or anti-CD3 stimulation. Additional studies showed that although PHA-stimulated HIV+ PBMC showed decreased median IL-2, IL-4, and TNF-alpha mRNA levels, they typically demonstrated more rapid kinetics (increased mean 4-h/24-h cytokine mRNA ratios), with significant differences for IL-4 (P<0.05) and TNF-alpha (P<0.005), compared to HIV- PBMC. The use of fresh or frozen cells gave comparable cytokine mRNA data; however, the secretion of some cytokine proteins (IL-2 receptor, IL-10, and TNF-alpha) appeared to be reduced in HIV+ PBMC that had been frozen and thawed. Our studies demonstrate that the kinetics of cytokine gene expression can reveal additional dysregulation of the immune system in HIV infection, suggesting that PBMC of HIV-infected persons exist in an activated state in vivo that permits them to express cytokine genes more rapidly than a normal PBMC.

Automatic poisson peak harvesting for high throughput protein identification.

High throughput identification of proteins by peptide mass fingerprinting requires an efficient means of picking peaks from mass spectra. Here, we report the development of a peak harvester to automatically pick monoisotopic peaks from spectra generated on matrix-assisted laser desorption/ionisation time of flight (MALDI-TOF) mass spectrometers. The peak harvester uses advanced mathematical morphology and watershed algorithms to first process spectra to stick representations. Subsequently, Poisson modelling is applied to determine which peak in an isotopically resolved group represents the monoisotopic mass of a peptide. We illustrate the features of the peak harvester with mass spectra of standard peptides, digests of gel-separated bovine serum albumin, and with Escherictia coli proteins prepared by two-dimensional polyacrylamide gel electrophoresis. In all cases, the peak harvester proved effective in its ability to pick similar monoisotopic peaks as an experienced human operator, and also proved effective in the identification of monoisotopic masses in cases where isotopic distributions of peptides were overlapping. The peak harvester can be operated in an interactive mode, or can be completely automated and linked through to peptide mass fingerprinting protein identification tools to achieve high throughput automated protein identification.