[The application and effect of bispherical augment in acetabular defects reconstruction].

Objective: To evaluate the clinical effect of bispherical augment in acetabular defects reconstruction in hip revision. Methods: This is a retrospective case series study. A retrospective analysis of 119 patients (124 hips) patients who underwent hip revision surgery and reconstructed with bispherical augment for acetabular bone defects from January 2019 to December 2023 was performed. There were 57 males (58 hips) and 62 females (66 hips), aged (65.0±11.8) years (range:40 to 102 years). The body mass index was (23.9±3.5) kg/m2 (range:16.1 to 32.2 kg/m2). Acetabular bone defects were typed as follows: 2 hips in Paprosky type ⅡA, 29 hips in type ⅡB, 34 hips in type ⅡC, 31 hips in type ⅢA, and 28 hips in type ⅢB, of which 9 patients (9 hips) were combined with pelvic discontinuity. Differences in Harris hip score (HHS) and lower limb discrepancy (LLD) were compared between preoperatively and final follow-up. The height of the hip center of rotation and the horizontal distance from the center of rotation to the teardrop were measured by radiographs before and after surgery, and prothesis stability and the occurrence of postoperative complications were evaluated. Data were compared using the paired sample t test. Results: All patients successfully completed the operation. The operation time was (167.0±53.4) minutes (range:90 to 380 minutes) and the intraoperative bleeding was (345.3±124.2) ml (range:100 to 1 200 ml). The height of the hip center of rotation decreased from (39.7±13.0) mm preoperatively to (21.8±7.1) mm postoperatively and the horizontal distance from the center of rotation to the teardrop increased from (34.0±10.1) mm preoperatively to (38.5±5.9) mm postoperatively, and the differences were statistically significant (t=15.859, P<0.01; t=5.266,P<0.01). All the patients were followed up for (26.1±15.4) months (range:6 to 60 months). At the last follow-up, HHS improved from (35.2±10.0) points preoperatively to (85.5±9.5) points, and the difference was statistically significant (t=50.723,P<0.01). LLD decreased from (2.1±1.1) cm preoperatively to (0.5±0.5) cm, and the difference was statistically significant (t=13.767, P<0.01). All acetabular components were stable and free of displacement on imaging during follow-up. Three patients suffered dislocation and received closed reduction, all prosthesis were in good position during follow-up. No dislocation, loosening, fracture, recurrence of infection and vascular nerve injury occurred in other patients. Conclusion: Bispherical augment can effectively reconstruct acetabular bone defects, restore the hip center of rotation, and improve hip joint function scores at short and mid-term follow-up.

[Efficacy comparison between a new generation of artificial ligaments and bone-patellar tendon-bone autograft for anterior cruciate ligament revision].

Objective: To compare the clinical efficacy of a new generation of ligaments (LARS artificial ligament) and bone-patellar tendon-bone (BPTB) autograft as grafts in anterior cruciate ligament (ACL) revision. Methods: A retrospective cohort study. The clinical data of 54 patients who underwent ACL revision from January 2018 to June 2020 in the First Hospital Affiliated to Army Medical University were retrospectively analyzed. There were 44 males and 10 females with a mean age of (28.5±7.7) years (15-45 years). Among them, 24 cases underwent ACL revision with LARS artificial ligament (LARS group), the other 30 cases underwent ACL revision with BPTB (BPTB group). The subjective and objective knee joint evaluation indexes were compared between the two groups to evaluate the clinical efficacy. The subjective evaluation indexes included Tegner score, Lysholm score and the International Knee Documentation Committee (IKDC) score. The objective evaluation indexes included the Lachman test, pivot-shift test, the anterior tibial translation (ATT) measurement at the weight-bearing position and the rate of patients returned to pre-injury sports. Results: The follow-up period was (32.8±5.3) months (24-42 months). At the last follow-up, the IKDC score, Tegner score and Lysholm score in the two groups significantly increased when compared with those before surgery (all P<0.05), and there was no significant difference in those indexes between the two groups (all P>0.05). The ATT measurement in the weight-bearing position was (3.1±0.7) mm in the LARS group and it was (4.1±0.9) mm in the BPTB group, which were significantly improved when compared with those before surgery (both P<0.05), and it was better in the LARS group than in the BPTB group (P<0.05). Postoperative Lachman test and pivot-shift test results in the LARS group were better than those in the BPTB group with statistically significant difference (both P<0.05). The rate of patients returned to pre-injury sports one year after surgery was 79.2%(19/24) in the LARS group and it was 50.0%(15/30) in the BPTB group, and the difference was statistically significant (P=0.029). Conclusions: Both LARS artificial ligament and BPTB autograft can achieve good short-term clinical efficacy in ACL revision, but LARS artificial ligament group has more advantages than BPTB autograft group in knee stability and early return to sports.

Optical limiting, nonlinear refraction and nonlinear absorption of the associates of Cd0.5Zn0.5S quantum dots and dyes.

The optical limiting properties of alloyed Cd0.5Zn0.5S quantum dots associated with erythrosine dye are analyzed using 532 nm, 40 ps pulses. We show that joint influence of saturable absorption, reverse saturable absorption and two-photon absorption cause the optical limiting of 532 nm radiation at the pulse energies exceeding 1 mJ. The nonlinear refraction and nonlinear absorption of these quantum dots associated with different organic dyes were studied using 1064 nm and 532 nm radiation. The nonlinear refraction index and nonlinear absorption coefficient of Cd0.5Zn0.5S quantum dots were measured at λ = 1064 nm to be 2 × 10-13 cm2 W-1 and 1.2 × 10-8 cm W-1, while the reverse saturable absorption of Cd0.5Zn0.5S quantum dots and erythrosine at λ = 532 nm was almost two orders larger. The potential applications of these quantum dots for high-order harmonic generation are discussed.

Upregulated microRNA-301a in osteosarcoma promotes tumor progression by targeting CDC14A.

MicroRNAs (miRs) are associated with tumor progression in various cancers, such as gastric and hepatic carcinomas, and lung cancer. miR-301a is overexpressed and displays oncogenic activity in cancers. We investigated the biological involvement of miR-301a in osteosarcoma (OS). Quantitative real-time polymerase chain reaction (qRT-PCR) was used to analyze expression levels of miR-301a in 24 OS and matched adjacent non-tumor tissues. A miR-301a mimic was transferred into OS cell lines U-2 OS and MG-63 to upregulate miR-301a. The effects of miR-301a were investigated by examining cell proliferation, migration, and the cell cycle. The miR-301 target was predicted by TargetScan and confirmed by western blotting and qRT-PCR. The expression of miR-301a was significantly higher in OS tissues compared with the matched adjacent non-tumor tissues (0.959 ± 0.39 vs 3.9516 ± 1.18). Upregulated miR-301a significantly increased proliferation at 48 and 72 h compared to the negative control (U-2 OS: 2.11 ± 0.21 vs 2.88 ± 0.24; 2.70 ± 0.26 vs 3.71 ± 0.24; MG-63: 2.19 ± 0.20 vs 3.19 ± 0.22; 3.1 ± 0.25 vs 4.01 ± 0.27) and migration capability (U-2 OS: 100 ± 20.19 vs 150.68 ± 32.83; MG-63: 100 ± 17.20 vs 133.35 ± 26.26), and decreased apoptosis in both U-2 OS (10.87 ± 2.53 vs 4.01 ± 2.23) and MG-63 (15.26 ± 2.15 vs 8.25 ± 3.07). The cell cycle studies revealed that miR-301a caused an increase of the G2 population in U-2 OS (38.6 ± 6.58 vs 47.2 ± 7.27) and MG-63 (44.01 ± 5.28 vs 57.9 ± 4.25). Additional experiments indicated that CDC14A was upregulated by miR-301a (0.63 ± 0.06 vs 0.98 ± 0.06; 1.49 ± 0.25 vs 2.99 ± 0.14). Overexpressed miR-301a may increase CDC14A expression and promote cell proliferation and migration in OS cells. Therefore, miR- 301a may be useful for osteosarcoma diagnosis and therapy.

Incidence of tardive dyskinesia: a comparison of long-acting injectable and oral paliperidone clinical trial databases.

BACKGROUND: To assess the tardive dyskinesia (TD) rate in studies of once-monthly long-acting injectable (LAI) paliperidone palmitate (PP) and once-daily oral paliperidone extended release (Pali ER). METHODS: Completed schizophrenia and bipolar studies for PP and Pali ER (≥ 6 month duration with retrievable patient-level data) were included in this post hoc analysis. Schooler-Kane research criteria were applied using Abnormal Involuntary Movement Scale (AIMS) scores to categorise probable (qualifying AIMS scores persisting for ≥ 3 months) and persistent TD (score persisting ≥ 6 months). Spontaneously reported TD adverse events (AEs) were also summarised. Impact of exposure duration on dyskinesia (defined as AIMS total score ≥ 3) was assessed by summarising the monthly dyskinesia rate. RESULTS: In the schizophrenia studies, TD rates for PP (four studies, N = 1689) vs. Pali ER (five studies, N = 2054), were: spontaneously reported AE, 0.18% (PP) vs. 0.10% (Pali ER); probable TD, 0.12% (PP) vs. 0.19% (Pali ER) and persistent TD, 0.12% (PP) vs. 0.05% (Pali ER). In the only bipolar study identified [Pali ER (N = 614)], TD rate was zero (spontaneously reported AE reporting, probable and persistent TD assessments). Dyskinesia rate was higher within the first month of treatment with both PP (13.1%) and Pali ER (11.7%) and steadily decreased over time (months 6-7: PP: 5.4%; Pali ER: 6.4%). Mean exposure: PP, 279.6 days; Pali ER, 187.2 days. CONCLUSIONS: Risk of TD with paliperidone was low (< 0.2%), regardless of the formulation (oral or LAI), in this clinical trial dataset. Longer cumulative exposure does not appear to increase the risk of dyskinesias.

Prediction of visual function from automatically quantified optical coherence tomography biomarkers in patients with geographic atrophy using machine learning.

Geographic atrophy (GA) is a vision-threatening manifestation of age-related macular degeneration (AMD), one of the leading causes of blindness globally. Objective, rapid, reliable, and scalable quantification of GA from optical coherence tomography (OCT) retinal scans is necessary for disease monitoring, prognostic research, and clinical endpoints for therapy development. Such automatically quantified biomarkers on OCT are likely to further elucidate structure-function correlation in GA and thus the pathophysiological mechanisms of disease development and progression. In this work, we aimed to predict visual function with machine-learning applied to automatically acquired quantitative imaging biomarkers in GA. A post-hoc analysis of data from a clinical trial and routine clinical care was conducted. A deep-learning automated segmentation model was applied on OCT scans from 476 eyes (325 patients) with GA. A separate machine learning prediction model (Random Forest) used the resultant quantitative OCT (qOCT) biomarkers to predict cross-sectional visual acuity under standard (VA) and low luminance (LLVA). The primary outcome was regression coefficient (r2) and mean absolute error (MAE) for cross-sectional VA and LLVA in Early Treatment Diabetic Retinopathy Study (ETDRS) letters. OCT parameters were predictive of VA (r2 0.40 MAE 11.7 ETDRS letters) and LLVA (r2 0.25 MAE 12.1). Normalised random forest feature importance, as a measure of the predictive value of the three constituent features of GA; retinal pigment epithelium (RPE)-loss, photoreceptor degeneration (PDR), hypertransmission and their locations, was reported both on voxel-level heatmaps and ETDRS-grid subfields. The foveal region (46.5%) and RPE-loss (31.1%) had greatest predictive importance for VA. For LLVA, however, non-foveal regions (74.5%) and PDR (38.9%) were most important. In conclusion, automated qOCT biomarkers demonstrate predictive significance for VA and LLVA in GA. LLVA is itself predictive of GA progression, implying that the predictive qOCT biomarkers provided by our model are also prognostic.

A strategy for rapid and efficient DNA sequencing by mass spectrometry.

Two methods of solid-phase Sanger DNA sequencing followed by detection with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry are demonstrated. In one method, sequencing ladders generated on an immobilized synthetic template were resolved up to the 63-mer including the primer. Detection sensitivity and resolution were sufficient for sequence analysis in the given range. This approach is particularly suitable for comparative (diagnostic) DNA sequencing. A second method that has the potential for high throughput de novo DNA sequencing is also presented; it uses immobilized duplex probes with five-base single-stranded overhangs to capture an unknown DNA template serving as primers for Sanger DNA sequencing. The power of mass spectrometry is demonstrated not only by its very high speed, but also by its ability to identify sequences that are not readable using gel electrophoresis.

Sequencing exons 5 to 8 of the p53 gene by MALDI-TOF mass spectrometry.

Matrix-assisted laser desorption ionization time of flight mass spectrometry was used to sequence exons 5 to 8 of the human p53 gene. A single tube procedure was established for target amplification and mass spectrometric (MS) sequencing. The MS sequencing scheme is designed for high throughput and parallel sample processing, and is amenable to full automation. Reliable sequencing data were obtained using fmol sample amounts. The high resolution and accuracy of MS sequencing was demonstrated by direct sequencing of a heterozygous template.

Estimated medical cost reductions for paliperidone palmitate vs placebo in a randomized, double-blind relapse-prevention trial of patients with schizoaffective disorder.

OBJECTIVE: The objective of this economic model was to estimate the difference in medical costs among patients treated with paliperidone palmitate once-monthly injectable antipsychotic (PP1M) vs placebo, based on clinical event rates reported in the 15-month randomized, double-blind, placebo-controlled, parallel-group study of paliperidone palmitate evaluating time to relapse in subjects with schizoaffective disorder. RESEARCH DESIGN AND METHODS: Rates of psychotic, depressive, and/or manic relapses and serious and non-serious treatment-emergent adverse events (TEAEs) were obtained from the long-term paliperidone palmitate vs placebo relapse prevention study. The total annual medical cost for a relapse from a US payer perspective was obtained from published literature and the costs for serious and non-serious TEAEs were based on Common Procedure Terminology codes. Total annual medical cost differences for patients treated with PP1M vs placebo were then estimated. Additionally, one-way and Monte Carlo sensitivity analyses were conducted. RESULTS: Lower rates of relapse (-18.3%) and serious TEAEs (-3.9%) were associated with use of PP1M vs placebo as reported in the long-term paliperidone palmitate vs placebo relapse prevention study. As a result of the reduction in these clinical event rates, the total annual medical cost was reduced by $7140 per patient treated with PP1M vs placebo. One-way sensitivity analysis showed that variations in relapse rates had the greatest impact on the estimated medical cost differences (range: -$9786, -$4670). Of the 10,000 random cycles of Monte Carlo simulations, 100% showed a medical cost difference <$0 (reduction) for patients using PPIM vs placebo. The average total annual medical differences per patient were -$8321 for PP1M monotherapy and -$6031 for PPIM adjunctive therapy. CONCLUSIONS: Use of PP1M for treatment of patients with schizoaffective disorder was associated with a significantly lower rate of relapse and a reduction in medical costs compared to placebo. Further evaluation in the real-world setting is warranted.

Lead poisoning: cause for alarm.

Lead (Pb) is everywhere in our environment and all children are exposed to lead to some degree. The Center for Disease Control (CDC) has recently declared that a blood lead level of 25 micrograms/dL and an erythrocyte protophoryrin of 35 micrograms/dL or above indicate an excessive absorption of lead in children and constitutes grounds for intervention. A study reported that about 780,000 (4%) of American children age 6 months to 5 years had blood lead levels of 30 micrograms/dL or higher. If the current level of 25 micrograms/dL had been used as a criterion in this study, the number of children involved would be much higher. It has been clearly stated by the American Academy of Pediatrics that ideally, all pre-school children should be screened for lead absorption by means of the erythrocyte protophoryrin test. The following is a review of lead poisoning in children and the serious consequences which may follow if it is not recognized early.

Lead poisoning in Hawaii: 1990.

Although lead (Pb) is one of the oldest known and most thoroughly described toxins, it continues to be a significant health hazard in 1990. There has been much progress in defining the nature and extent of low-level lead toxicity during the past decade. There continues to be insidious sources of lead toxicity in our environment, in water, food, paint and contaminated soil. As the epidemiology of lead poisoning is more clearly defined, toxicities are recognized as the result of lower and lower levels of exposure. Recognition of low-level lead exposure and the primary prevention of its effects on health requires a keen awareness of high-risk environments as well as the subtle symptoms and signs of lead poisoning. A high index of suspicion by primary care physicians plus government support are necessary to implement successful prevention programs.

An early Cambrian agglutinated tubular lophophorate with brachiopod characters.

The morphological disparity of lophotrochozoan phyla makes it difficult to predict the morphology of the last common ancestor. Only fossils of stem groups can help discover the morphological transitions that occurred along the roots of these phyla. Here, we describe a tubular fossil Yuganotheca elegans gen. et sp. nov. from the Cambrian (Stage 3) Chengjiang Lagerstätte (Yunnan, China) that exhibits an unusual combination of phoronid, brachiopod and tommotiid (Cambrian problematica) characters, notably a pair of agglutinated valves, enclosing a horseshoe-shaped lophophore, supported by a lower bipartite tubular attachment structure with a long pedicle with coelomic space. The terminal bulb of the pedicle provided anchorage in soft sediment. The discovery has important implications for the early evolution of lophotrochozoans, suggesting rooting of brachiopods into the sessile lophotrochozoans and the origination of their bivalved bauplan preceding the biomineralization of shell valves in crown brachiopods.

Importance of specific adenosine N7-nitrogens for efficient cleavage by a hammerhead ribozyme. A model for magnesium binding.

Five modified hammerhead ribozyme/substrate complexes have been prepared in which individual adenosine N7-nitrogens have been excised. The modified complexes were chemically synthesized with the substitution of a single 7-deazaadenosine (c7A) base analogue for residues A11, A14, A26, A27, or A28. Two of the base analogues, c7A11 and c7A14, occur in a 19-mer ribozyme, while the remaining three residues, c7A26, c7A27, and c7A28, are present in a 24-mer substrate. Under stoichiometric conditions, four of the complexes are cleaved with relatively little change in rate when compared with that of the native complex. However, the relative rate for the c7A11 complex is some 35-fold slower than that of the native complex. Steady-state kinetic analyses indicate that the cleavage efficiencies, as measured by kcat/KM, for the c7A14, c7A26, c7A27, and c7A28 complexes are reduced 18-fold, 10-fold, 34-fold, and 16-fold, respectively. These reductions in cleavage efficiency are primarily a result of lower kcat values. By comparison, the cleavage efficiency of the c7A11 complex is reduced more than 200-fold relative to that of the native complex, again primarily as a result of a lower kcat value. The results suggest that the N7-nitrogen of A11 in the hammerhead ribozyme/substrate complex is critical for efficient cleavage activity. The results of the present work, in combination with those from previous reports, indicate that five critical functional groups are located within the tetrameric sequence G10A11U12G13. A preliminary model for the binding of a single magnesium cofactor to this portion of the sequence is proposed.(ABSTRACT TRUNCATED AT 250 WORDS)

Importance of specific purine amino and hydroxyl groups for efficient cleavage by a hammerhead ribozyme.

Eight modified ribozymes of 19 residues have been prepared with individual purine amino or hydroxyl groups excised. The modified ribozymes were chemically synthesized with the substitution of a single 2'-deoxyadenosine, 2'-deoxyguanosine, inosine, or purine riboside for residues G10, A11, G13, or A14. Five of the modified ribozymes cleaved the 24-mer substrate with little change in rate as monitored by simple first-order kinetics. However, deletion of the 2-amino group at G10 (replacement with inosine) or deletion of either of the 2'-hydroxyls at G10 or G13 (replacement with 2'-deoxyguanosine) resulted in ribozymes with a drastic decrease in cleavage efficiency. Increasing the concentration of the Mg2+ cofactor from 10 mM to 50 mM significantly enhanced cleavage efficiency by these three derivatives. Steady-state kinetic assays for these three ribozymes indicated that the modifications result in both an increase in Km and a decrease in kcat. These results suggest that the exocyclic amino group at-G10 and the hydroxyls at G10 and G13 are important both for ribozyme-substrate binding and for the Mg(2+)-catalyzed cleavage reaction.

Importance of specific guanosine N7-nitrogens and purine amino groups for efficient cleavage by a hammerhead ribozyme.

Seven modified hammerhead ribozyme/substrate complexes have been prepared in which individual purine nitrogens, the guanine N7-, the guanine N2-, or the adenine N6-nitrogen, have been excised. The modified complexes were chemically synthesized with the substitution of a single 7-deazaguanosine (c7G), inosine (I), or nebularine (purine riboside, P) base analogue as appropriate for residues G5, G8, G12, A13, A14, or A15. Two of the base analogues, c7G5 and C7G8, occur in a 19-mer ribozyme, while the remaining three residues are present in a 24-mer substrate. Under stoichiometric conditions, four of the complexes, G5c7G, G8c7G, G12c7G, and A14P, are cleaved with relatively little change in rate when compared with the native complex. Two complexes, A13P and A15P, are cleaved some 6-8-fold slower than the native complex, while the G12I complex is reduced in rate by 50-fold. Steady-state kinetic analyses indicate that the cleavage efficiencies, as measured by kcat/KM values, for the G5c7G, G8c7G, and G12c7G complexes are only marginally reduced relative to the native complex. The values for the A13P, A14P, and A15P complexes are reduced by 25-, 15-, and 60-fold, respectively. These reductions in cleavage efficiency are primarily a result of lower kcat values. By comparison, the kcat/KM value for the G12I complex is decreased 450-fold relative to the native complex and is characterized by an 8-fold increase in KM and a kcat value that is reduced nearly 60-fold. These results indicate that the N2-amino group of G12 in the hammerhead ribozyme/substrate complex is critical for efficient cleavage activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Activity of the hammerhead ribozyme upon inversion of the stereocenters for the guanosine 2'-hydroxyls.

Two guanosine 2'-hydroxyls in the hammerhead RNA complex at positions G5 and G8 are critical for efficient cleavage by this RNA catalyst. These two functional groups are likely involved in the binding of the metal cofactor, or they are involved in specific interresidue hydrogen-bonding interactions. The importance of the stereochemical positioning of both critical 2'-hydroxyls was investigated by comparing the cleavage rates of three arabinosylguanine-substituted complexes (in which the positions of specific guanosine 2'-hydroxyls were stereochemically altered by inverting the C2' stereocenter) with that of the native complex, as well as with the rates of the dG- and dFG-substituted complexes [in which the 2'-hydroxyls are absent as the result of substitution by 2'-deoxyguanosine (dG) or 2'-deoxy-2'-fluoroguanosine (dFG)]. The G5araG and G8araG complexes exhibit dramatically different cleavage rates. The G5araG complex is essentially inactive, at least 10(5)-fold slower than the native complex. RNA cleavage by this analogue ribozyme is also 1000-fold slower than cleavage by either the G5dG or the G5dFG ribozyme, both of which lack the 2'-hydroxyl at G5. By comparison, catlytic efficiency of the G8araG complex as expressed by kcat/Km is comparable with that of the native complex and some 2 orders of magnitude more active than either the G8dG or the G8dFG complex.

Efficient preparation of short DNA sequence ladders potentially suitable for MALDI-TOF DNA sequencing.

Duplex probes with five-base single-stranded overhangs were developed for positional sequencing by hybridization [Broude et al., Proc Natl Acad Sci USA 91:3072-3076, 1994]. The partially duplex probes can be employed to capture single-stranded oligonucleotide targets and form primer-template complexes. Recently we showed that partially duplex probes can prime Sanger sequencing reactions on immobilized, but non-ligated long single-stranded targets (approximately 500 nucleotide) [Fu et al., Proc Natl Acad Sci, in press]. Here immobilized, non-ligated partially duplex probes were used to capture and sequence short single-stranded targets. This strategy is capable of rapidly preparing large numbers of samples for future mass spectrometric DNA sequencing.

Sequencing double-stranded DNA by strand displacement.

Duplex probes with five base single-stranded overhangs can capture dsDNA targets from type IIS restriction nuclease digests. Ligation generates a predesigned nick site, where DNA polymerase can generate sequencing ladders by strand displacement or nick translation in the presence of trace amounts of dideoxynucleotides. This allows dsDNA targets to be captured from mixtures and directly sequenced without subcloning, purification or denaturation.

A DNA sequencing strategy that requires only five bases of known terminal sequence for priming.

We have previously reported an enhanced version of sequencing by hybridization (SBH), termed positional SBH (PSBH). PSBH uses partially duplex probes containing single-stranded 3' overhangs, instead of simple single-stranded probes. Stacking interactions between the duplex probe and a single-stranded target allow us to reduce the probe sizes required to 5-base single-stranded overhangs. Here we demonstrate the use of PSBH to capture relatively long single-stranded DNA targets and perform standard solid-state Sanger sequencing on these primer-template complexes without ligation. Our results indicate that only 5 bases of known terminal sequence are required for priming. In addition, the partially duplex probes have the ability to capture their specific target from a mixture of five single-stranded targets with different 3'-terminal sequences. This indicates the potential utility of the PSBH approach to sequence mixtures of DNA targets without prior purification.

Chip-based genotyping by mass spectrometry.

Silicon chips with immobilized target DNAs were used for accurate genotyping by mass spectrometry. Genomic DNAs were amplified with PCR, and the amplified products were covalently attached to chip wells via N-succinimidyl (4-iodoacetyl)aminobenzoate (SIAB) chemistry. Primer annealing, extension, and termination were performed on a 1-microl scale directly in the chip wells in parallel. Diagnostic products thus generated were detected in situ by using matrix-assisted laser desorption ionization mass spectrometry. This miniaturized method has the potential for accurate, high-throughput, low-cost identification of genetic variations.