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Veno-veous extracorporeal membrane oxygenation (VV-ECMO) has been widely used in the treatment for severe acute respiratory distress syndrome (ARDS). Up to now, the routine access to establish VV-ECMO involves two-sites single lumen cannula via femoral vein and internal jugular venous in adult and children, while few studies about the dual lumen cannula (DLC) in VV-ECMO implemented in adult and children have been reported. On December 16, 2021, an unconscious child with severe ARDS due to multiple trauma caused by fatal falling from a height was admitted to Taihe Hospital. The initial diagnosis was hemorrhagic shock, bilateral hemopneumothorax, sternal fracture, cavity organ perforation, splenic rupture, and pelvic fracture and severe ARDS. Despite mechanical ventilation, he progressed to refractory hypoxemia and was treated with VV-ECMO after successful DLC placement in the right internal jugular vein by the mobile ECMO team of intensive care unit of the Union Hospital eventually. In addition, he received endoscopic sputum aspiration, prone position ventilation, anti-infection and nutritional treatment. His oxygenation gradually improved and he was successfully weaned from ECMO after 11 days. In this case, DLC simplified the process without any related complications, suggesting that it can be safely and effectively used in the treatment of Child's severe ARDS.

Convalescent plasma therapy has been implemented in a few cases of severe coronavirus disease 2019. No report about convalescent plasma therapy in treating patients with prolonged positivity of SARS-CoV-2 RNA has been published. In this study, we conducted a retrospective observational study in 27 patients with prolonged positivity of SARS-CoV-2 RNA, the clinical benefit of convalescent plasma therapy were analyzed. qRT-PCR test of SARS-CoV-2 RNA turned negative (≤ 7 days) in a part of patients (early negative group, n = 15) after therapy, others (late negative group, n = 12) turned negative in more than 7 days. Pulmonary imaging improvement was confirmed in 7 patients in early negative group and 8 in late negative group after CP therapy. Viral load decreased in early negative group compared with late negative group at day 3, 5, 7 after implementing convalescent plasma therapy. Patients in early negative group had a shorter median length of hospital stay. In conclusion, convalescent plasma therapy might help eliminate virus and shorten length of hospital stay in patients with prolonged positivity of SARS-CoV-2 RNA.

The coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, has spread around the world with high mortality. To diagnose promptly and accurately is the vital step to effectively control its pandemic. Dynamic characteristics of SARS-CoV-2-specific antibodies which are important for diagnosis of infection have not been fully demonstrated. In this retrospective, single-center, observational study, we enrolled the initial 131 confirmed cases of COVID-19 at Jin-Yin-Tan Hospital who had at least one-time antibody tested during their hospitalization. The dynamic changes of IgM and IgG antibodies to SARS-CoV-2 nucleocapsid protein in 226 serum samples were detected by ELISA. The sensitivities of IgM and IgG ELISA detection were analyzed. Result showed that the sensitivity of the IgG ELISA detection (92.5%) was significantly higher than that of the IgM (70.8%) (P

Background A COVID-19 outbreak started in Wuhan, China, last December and now has become a global pandemic. The clinical information in caring of critically ill patients with COVID-19 needs to be shared timely, especially under the situations that there is still a largely ongoing spread of COVID-19 in many countries. Methods A multicenter prospective observational study investigated all the COVID-19 patients received in 19 ICUs of 16 hospitals in Wuhan, China, over 24 h between 8 AM February 2h and 8 AM February 27, 2020. The demographic information, clinical characteristics, vital signs, complications, laboratory values, and clinical managements of the patients were studied. Results A total of 226 patients were included. Their median (interquartile range, IQR) age was 64 (57–70) years, and 139 (61.5%) patients were male. The duration from the date of ICU admission to the study date was 11 (5–17) days, and the duration from onset of symptoms to the study date was 31 (24–36) days. Among all the patients, 155 (68.6%) had at least one coexisting disease, and their sequential organ failure assessment score was 4 (2–8). Organ function damages were found in most of the patients: ARDS in 161 (71.2%) patients, septic shock in 34 (15.0%) patients, acute kidney injury occurred in 57 (25.2%) patients, cardiac injury in 61 (27.0%) patients, and lymphocytopenia in 160 (70.8%) patients. Of all the studied patients, 85 (37.6%) received invasive mechanical ventilation, including 14 (6.2%) treated with extracorporeal membrane oxygenation (ECMO) at the same time, 20 (8.8%) received noninvasive mechanical ventilation, and 24 (10.6%) received continuous renal replacement therapy. By April 9, 2020, 87 (38.5%) patients were deceased and 15 (6.7%) were still in the hospital. Conclusions Critically ill patients with COVID-19 are associated with a higher risk of severe complications and need to receive an intensive level of treatments. COVID-19 poses a great strain on critical care resources in hospitals. Trial registration Chinese Clinical Trial Registry, ChiCTR2000030164. Registered on February 24, 2020, http://www.chictr.org.cn/edit.aspx?pid=49983&htm=4

BackgroundDifferentiated thyroid cancer (DTC) is usually very treatable and is often cured with surgery and, if indicated, follow with radioactive iodine (RAI) and long-term thyroid-stimulating hormone (TSH) suppression with levothyroxine. Unfortunately, locoregional relapse and distant metastases occur in up to 20% at ten years; those relapsed or distant metastatic thyroid cancer is relatively uncommon compare to other cancers, cytotoxic chemotherapies provided low response rates, and two-thirds of cases became RAI refractory (RAI-R). New therapy options for local-regional relapsed or distant metastases thyroid cancer are desperately needed. Chimeric antigen receptor T cells (CAR-T) have been demonstrated remarkable efficacy in hematological cancers but have not yet translated in treating solid tumors. The significant hurdles limiting CAR-T therapy were due to a paucity of differentially expressed cell surface molecules on solid tumors that can be safely targeted. Here, we present thyroid-stimulating hormone receptor (TSHR) as a putative target for CAR-T therapy of DTC.MethodsWe undertook a large-scale screen on thyroid cancer tissues and multiple internal organs through bioinformatical analysis and immunohistochemistry to date TSHR expression. Using three previously described mAb, we generate three third-generation CAR-Ts. We tested anti-TSHR CAR-T in vitro activity by T-cell function assay and killing assay. Then we tested pre-clinical therapeutical efficacy in a xenograft mouse model of DTC and analyzed mice’s physical conditions and histological abnormalities to evaluate anti-TSHR CAR-T’s safety.ResultsTSHR is highly and homogeneously expressed on 90.8%(138/152) of papillary thyroid cancer, 89.2% (33/37) of follicular thyroid cancer, 78.2% (18/23) of the cervical lymph node metastases, and 86.7% of locally recurrent or metastasis lesions with RAI-R disease. We develop three novel anti-TSHR CAR-T from mAb M22, K1-18, and K1-70, all three CAR-Ts mediate significant anti-tumor activity in vitro. Among these, we demonstrate that K1-70 CAR-T can have therapeutical efficacy in vivo, and no apparent toxicity has been observed.ConclusionTSHR is a latent target antigen of CAR-T therapy for DTC. Anti-TSHR CAR-T has strong therapeutical efficacy in vivo while without prominent adverse events. Anti-TSHR CAR-T could represent an exciting therapeutic option for patients with specific local-regional relapsed or distant metastases thyroid cancer and should be tested in carefully designed clinical trials.

Background: The outbreak of coronavirus disease 2019 (COVID-19) has led to a large and increasing number of patients requiring prolonged mechanical ventilation and tracheostomy. The indication and optimal timing of tracheostomy in COVID-19 patients are still unclear, and the outcomes about tracheostomy have not been extensively reported. We aimed to describe the clinical characteristics and outcomes of patients with confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pneumonia who underwent elective tracheostomies.Methods: The multi-center, retrospective, observational study investigated all the COVID-19 patients who underwent elective tracheostomies in intensive care units (ICUs) of 23 hospitals in Hubei province, China, from January 8, 2020 to March 25, 2020. Demographic information, clinical characteristics, treatment, details of the tracheostomy procedure, successful weaning after tracheostomy, and living status were collected and analyzed. Data were compared between early tracheostomy patients (tracheostomy performed within 14 days of intubation) and late tracheostomy patients (tracheostomy performed after 14 days).Results: A total of 80 patients were included. The median duration from endotracheal intubation to tracheostomy was 17.5 [IQR 11.3–27.0] days. Most tracheotomies were performed by ICU physician [62 (77.5%)], and using percutaneous techniques [63 (78.8%)] at the ICU bedside [76 (95.0%)]. The most common complication was tracheostoma bleeding [14 (17.5%)], and major bleeding occurred in 4 (5.0%) patients. At 60 days after intubation, 31 (38.8%) patients experienced successful weaning from ventilator, 17 (21.2%) patients discharged from ICU, and 43 (53.8%) patients had died. Higher 60 day mortality [22 (73.3%) vs. 21 (42.0%)] were identified in patients who underwent early tracheostomy.Conclusions: In patients with SARS-CoV-2 pneumonia, tracheostomies were feasible to conduct by ICU physician at bedside with few major complications. Compared with tracheostomies conducted after 14 days of intubation, tracheostomies within 14 days were associated with an increased mortality rate.

Due to type-setting error, the affiliation of author Huaqing Shu and Shuzhen Wang was mislabeled.

Background The global numbers of confirmed cases and deceased critically ill patients with COVID-19 are increasing. However, the clinical course, and the 60-day mortality and its predictors in critically ill patients have not been fully elucidated. The aim of this study is to identify the clinical course, and 60-day mortality and its predictors in critically ill patients with COVID-19. Methods Critically ill adult patients admitted to intensive care units (ICUs) from 3 hospitals in Wuhan, China, were included. Data on demographic information, preexisting comorbidities, laboratory findings at ICU admission, treatments, clinical outcomes, and results of SARS-CoV-2 RNA tests and of serum SARS-CoV-2 IgM were collected including the duration between symptom onset and negative conversion of SARS-CoV-2 RNA. Results Of 1748 patients with COVID-19, 239 (13.7%) critically ill patients were included. Complications included acute respiratory distress syndrome (ARDS) in 164 (68.6%) patients, coagulopathy in 150 (62.7%) patients, acute cardiac injury in 103 (43.1%) patients, and acute kidney injury (AKI) in 119 (49.8%) patients, which occurred 15.5 days, 17 days, 18.5 days, and 19 days after the symptom onset, respectively. The median duration of the negative conversion of SARS-CoV-2 RNA was 30 (range 6–81) days in 49 critically ill survivors that were identified. A total of 147 (61.5%) patients deceased by 60 days after ICU admission. The median duration between ICU admission and decease was 12 (range 3–36). Cox proportional-hazards regression analysis revealed that age older than 65 years, thrombocytopenia at ICU admission, ARDS, and AKI independently predicted the 60-day mortality. Conclusions Severe complications are common and the 60-day mortality of critically ill patients with COVID-19 is considerably high. The duration of the negative conversion of SARS-CoV-2 RNA and its association with the severity of critically ill patients with COVID-19 should be seriously considered and further studied.

Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes novel coronavirus disease 2019 (COVID-19), is spreading rapidly around the world. Thrombocytopenia in patients with COVID-19 has not been fully studied. Objective To describe thrombocytopenia in patients with COVID-19. Methods For each of 1476 consecutive patients with COVID-19 from Jinyintan Hospital, Wuhan, China, nadir platelet count during hospitalization was retrospectively collected and categorized into (0, 50], (50, 100], (100-150], or (150-) groups after taking the unit (×109 /L) away from the report of nadir platelet count. Nadir platelet counts and in-hospital mortality were analyzed. Results Among all patients, 238 (16.1%) patients were deceased and 306 (20.7%) had thrombocytopenia. Compared with survivors, non-survivors were older, were more likely to have thrombocytopenia, and had lower nadir platelet counts. The in-hospital mortality was 92.1%, 61.2%, 17.5%, and 4.7% for (0, 50], (50, 100], (100-150], and (150-) groups, respectively. With (150-) as the reference, nadir platelet counts of (100-150], (50, 100], and (0, 50] groups had a relative risk of 3.42 (95% confidence interval [CI] 2.36-4.96), 9.99 (95% CI 7.16-13.94), and 13.68 (95% CI 9.89-18.92), respectively. Conclusions Thrombocytopenia is common in patients with COVID-19, and it is associated with increased risk of in-hospital mortality. The lower the platelet count, the higher the mortality becomes.

Summary Background An ongoing outbreak of pneumonia associated with the severe acute respiratory coronavirus 2 (SARS-CoV-2) started in December, 2019, in Wuhan, China. Information about critically ill patients with SARS-CoV-2 infection is scarce. We aimed to describe the clinical course and outcomes of critically ill patients with SARS-CoV-2 pneumonia. Methods In this single-centered, retrospective, observational study, we enrolled 52 critically ill adult patients with SARS-CoV-2 pneumonia who were admitted to the intensive care unit (ICU) of Wuhan Jin Yin-tan hospital (Wuhan, China) between late December, 2019, and Jan 26, 2020. Demographic data, symptoms, laboratory values, comorbidities, treatments, and clinical outcomes were all collected. Data were compared between survivors and non-survivors. The primary outcome was 28-day mortality, as of Feb 9, 2020. Secondary outcomes included incidence of SARS-CoV-2-related acute respiratory distress syndrome (ARDS) and the proportion of patients requiring mechanical ventilation. Findings Of 710 patients with SARS-CoV-2 pneumonia, 52 critically ill adult patients were included. The mean age of the 52 patients was 59·7 (SD 13·3) years, 35 (67%) were men, 21 (40%) had chronic illness, 51 (98%) had fever. 32 (61·5%) patients had died at 28 days, and the median duration from admission to the intensive care unit (ICU) to death was 7 (IQR 3–11) days for non-survivors. Compared with survivors, non-survivors were older (64·6 years [11·2] vs 51·9 years [12·9]), more likely to develop ARDS (26 [81%] patients vs 9 [45%] patients), and more likely to receive mechanical ventilation (30 [94%] patients vs 7 [35%] patients), either invasively or non-invasively. Most patients had organ function damage, including 35 (67%) with ARDS, 15 (29%) with acute kidney injury, 12 (23%) with cardiac injury, 15 (29%) with liver dysfunction, and one (2%) with pneumothorax. 37 (71%) patients required mechanical ventilation. Hospital-acquired infection occurred in seven (13·5%) patients. Interpretation The mortality of critically ill patients with SARS-CoV-2 pneumonia is considerable. The survival time of the non-survivors is likely to be within 1–2 weeks after ICU admission. Older patients (>65 years) with comorbidities and ARDS are at increased risk of death. The severity of SARS-CoV-2 pneumonia poses great strain on critical care resources in hospitals, especially if they are not adequately staffed or resourced. Funding None.

The pandemic of the coronavirus disease 2019 (COVID-19) has become a global public health crisis. The symptoms of COVID-19 range from mild to severe, but the physiological changes associated with COVID-19 are barely understood. In this study, we performed targeted metabolomic and lipidomic analyses of plasma from a cohort of patients with COVID-19 who had experienced different symptoms. We found that metabolite and lipid alterations exhibit apparent correlation with the course of disease in these patients, indicating that the development of COVID-19 affected their whole-body metabolism. In particular, malic acid of the TCA cycle and carbamoyl phosphate of the urea cycle result in altered energy metabolism and hepatic dysfunction, respectively. It should be noted that carbamoyl phosphate is profoundly down-regulated in patients who died compared with patients with mild symptoms. And, more importantly, guanosine monophosphate (GMP), which is mediated not only by GMP synthase but also by CD39 and CD73, is significantly changed between healthy subjects and patients with COVID-19, as well as between the mild and fatal cases. In addition, dyslipidemia was observed in patients with COVID-19. Overall, the disturbed metabolic patterns have been found to align with the progress and severity of COVID-19. This work provides valuable knowledge about plasma biomarkers associated with COVID-19 and potential therapeutic targets, as well as an important resource for further studies of the pathogenesis of COVID-19.

Additional file 1: S1. Symptoms of 239 critically ill patients with COVID-19.

To investigate the influence of heparin pretreatment on serum and lung tissue level of neutrophil extracellular traps (NETs) in septic mice model and its molecular mechanism.Ninety male C57BL/6J mice were randomly divided into control group (n = 30), lipopolysaccharides (LPS) group (n = 30, 30 mg/kg LPS in 100 ?L normal saline was intraperitoneally injected) and LPS+heparin group (n = 30, 8 U of heparin in 20 ?L normal saline was subcutaneously injected 30 minutes before the injection of LPS). Six hours later of LPS injection, blood was collected and lung tissue was harvested. Enzyme linked immunosorbent assay (ELISA) was used to assess the concentration of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and histones 2AX (H2AX), neutrophil elastase (NE), which reflected NETs concentration. PicoGreen fluorescent dyes was used to detect serum circulating free DNA (cf-DNA/NETs) concentration. The protein expression levels of H2AX and NE in lung tissue were examined by Western Blot.The serum concentrations of TNF-α, IL-6, H2AX, NE, cf-DNA/NETs, and the protein expression levels of H2AX and NE in lung tissue of septic mice were significantly higher than those of control group [TNF-α (ng/L): 133.0±14.1 vs. 2.7±1.0, IL-6 (ng/L): 3 911.2±189.2 vs. 298.9±52.5, H2AX (ng/L): 545.5±40.0 vs. 21.9±8.3, NE (μg/L): 6.48±0.12 vs. 0.47±0.15, cf-DNA/NETs (μg/L): 846.3±137.5 vs. 152.7±36.4, H2AX protein (gray value): 1.14±0.09 vs. 0.68±0.04, NE protein (gray value): 0.56±0.03 vs. 0.32±0.04, all P0.05]. After heparin pretreatment, levels of serum TNF-α, H2AX, NE, cf-DNA/NETs, and protein expression levels of H2AX and NE in lung tissue were significantly reduced [TNF-α (ng/L): 83.2±7.6 vs. 133.0±14.1, H2AX (ng/L): 435.0±39.0 vs. 545.5±40.0, NE (μg/L): 4.26±0.17 vs. 6.48±0.12, cf-DNA/NETs (μg/L): 606.5±73.9 vs. 846.3±137.5, H2AX protein (gray value): 0.91±0.03 vs. 1.14±0.09, NE protein (gray value): 0.42±0.03 vs. 0.56±0.03, all P0.05], but no significant change was found in IL-6 (ng/L: 3 919.9±166.6 vs. 3 911.2±189.2, P0.05).Heparin pretreatment could significantly decrease the level of NETs in serum and lung tissue, and can be the potential mechanism of its organ protection in sepsis.

Magnetizations of the Fe doped Sr 3 Cr 2 O 8 single crystals Sr 3 Cr 2− x Fe x O 8 ( x =0, 0.05 and 0.1) were investigated in the temperature ranging from 2 to 300 K and the magnetic field ranging from 0 to 16 T. Based on the Curie–Weiss law and the Bleaney–Bowers model, the low field magnetization behaviors were analyzed. Experimental results show that the Fe 5+ dopant has influence on the intradimer interaction J 0 , the interdimer interaction J' , and the average change rate Δ χ / Δ T of spin-dimer susceptibility χ d i m e r of Sr 3 Cr 2− x Fe x O 8 . Especially, in this spin-dimer system, the Fe 5+ dopant can obviously enhance the magnetic sensitivity of singlet to triplet transition in Sr 3 Cr 2 O 8 .

Measurements of the isothermal magnetization on La0.27Nd0.4Ca0.33MnO3 have been performed in a field up to 7 T at various temperatures. A distinct field-induced metamagnetic transition is observed within a temperature region of 2–180 K, resulting from the coexistence of charge ordered (CO), antiferromagnetic (AFM) and ferromagnetic (FM) phases. A temperature and magnetic field dependent irreversible CO–FM transition appears in the lower temperature region. Accompanying the transition an obvious lattice strain was observed, indicating that the transition is related to the change of lattice strain due to strong spin–lattice coupling in La0.27Nd0.4Ca0.33MnO3. These properties make the doped manganites a good candidate for a magnetostriction sensor material.

High-quality single-crystal Sr6Co5O15–δ (δ = 1.33) was successfully synthesized with a flux method. Detailed crystalline structure was determined by single-crystal X-ray diffraction at 298 K, which revealed a quasi-one-dimensional Co–O chain-like structure of this oxygen-deficient material. The magnetizations were measured in a pulsed high magnetic field up to 32 T and with a Physical Property Measurement System (PPMS). A large anisotropy and a distinct spin-flip transition were observed in the magnetization behavior. Theoretical calculation based on Botana's results suggests that interchain interaction (non-one-dimensional magnetic coupling) plays an important role in the magnetic properties of Sr6Co5O15–δ.

We have synthesized compounds of Ca3Co2−xFexO6 (x = 0–1.5) by sol–gel reaction method. X-ray diffraction data show that the solubility limit of Fe in the spin-chain compound Ca3Co2O6 is about x = 0.7. For the rhombohedral ( R 3 ¯ c ) samples with x ≤ 0.7, the upward deviation from the paramagnetic Curie–Weiss law in the inverse magnetic susceptibility (H/M) tends to be suppressed at x = 0.7. Correlatively, the paramagnetic Curie temperature θP approaches zero at this concentration. The M(H) curves exhibit a well-defined ferrimagnetic state, which becomes smeared out with increasing Fe concentration and nearly disappears at x = 0.7. For the triclinic ( P 1 ¯ ) samples with x = 1.1, the magnetization is much small and its ground state is antiferromagnetic. The H/M curve measured up to 300 K does not follow the Curie–Weiss law. Finally, ESR spectra composed of eight hyperfine splittings due to Co nuclear spin are observed for x = 0.4.

The electrical transport and magnetization measurements have been carried out on Al-doped polycrystalline intermetallic compounds Dy 50− x Al x Ag 50 ( x = 0, 0.3, 0.6, 1.2, 1.8) in a pulsed high magnetic field, in which multi-step magnetization is observed. Partial substitute of non-magnetic Al 3+ for Dy 3+ ions in the compounds increases the critical magnetic fields and the relative area of the magnetic hysteresis loop, which result from the pinning effect, lattice distortion, the change of coupling strength and dilution effect related to the Al 3+ doping.The experimental results indicate that non-magnetic Al 3+ ions and induced amorphous phase can pin the rotation and/or growth of magnetic domains, thus, the critical magnetic field can be enhanced by doping non-magnetic ions in the magnetic materials, especially in the permanent magnet materials.

Magnetization behavior of (La 0.83 Bi 0.17 ) 0.67 Ca 0.33 MnO 3 has been investigated in the temperature range from 100 to 180 K. A metamagnetic transition was observed in the temperature region, where the magnetization was measured after a zero-field-cooling from room temperature to a selected temperature. Experimental results show that, after a higher magnetization route, the field-increasing branches of the magnetization curves shows an unusual training effect: below a magnetic field H 0 , the applied magnetic field enhances the value of magnetization; however, above H 0 the magnetic field suppresses the value, and the behavior cannot be totally attributed to the enhancement effect of the applied magnetic field on ferromagnetic phase fraction. It is proposed that, in the two-phase coexistence region, the higher magnetic field promotes the phase separation and leads to both the fraction of ferromagnetic domain and the stabilization of antiferromagnetic domain increase.

Summary Background Venous thrombosis (VT) is a worldwide medical problem. In order to identify individuals at high risk early, it is necessary to find more genetic risk factors. Nowadays, the studies on genetic factors of thrombosis are mainly focused on coagulation and anticoagulation factors. The exploration of other proteins involved in thrombosis and hemostasis may lead to a breakthrough. Objectives We used APOH as a candidate gene to investigate the existence of genetic variation that could increase the risk of thrombosis. Methods/Results In the current study, with a resequencing method followed by a case–control study, four polymorphisms (c.−32C>A, c.422T>C, c.461G>A, and c.1004G>C) in APOH (encoding β2-glycoprotein I) were found to be in high linkage disequilibrium, which could result in three haplotypes. The H2 heterozygotes and H3 homozygotes had approximately 1.5-fold and seven-fold increased risks for VT, respectively. The minor allele frequency in the general population was ~ 10%. In addition, H3 individuals showed a significantly decreased level of β2-glycoprotein I, but an increased level of thrombin generation. Functional tests indicated that the mutant β2-glycoprotein I had a significantly lower capacity to extend thrombin clotting time and increase thrombin generation potential. Conclusions This study revealed APOH as a new candidate gene associated with thrombosis, and further genetic research on this gene in patients in whom the cause of thrombophilia is unknown is therefore warranted.

Transport measurements of as-grown and annealed (Ga,Mn)As samples were conducted under pulsed high magnetic field up to 40T. Assuming that the magnetization follows modified Brillouin function, we obtained the hole densities of the (Ga,Mn)As samples at various temperatures, which generally increase with temperature and become two times larger after annealing process. The hole density determined by electrical transport measurement at low magnetic field up to 1.5T is found to be underestimated.