Tape-stripped Skin Research Articles

Rapid analysis of tetracaine for a tape stripping pharmacokinetic study using short‐end capillary electrophoresis

A rapid and simple short-end (reverse) capillary zone electrophoresis method was developed and validated for the separation and quantification of tetracaine in skin using tape samples. The separation was performed in a 485 mm (400 mm to window) x 50 microm internal diameter fused silica capillary using a background electrolyte of phosphoric acid-Tris pH2.5 at -25 kV. The extraction of tetracaine from tape samples was achieved using methanol diluted to 50% with water before injection. Procaine was the internal standard. The migration times for procaine and tetracaine were 1.25 and 1.36 min, respectively. The limit of quantification for tetracaine was 50 microg, with a signal-to-noise ratio greater than 10. The calibration curve was linear from 50 to 1200 microg with r(2) greater than 0.99. The CV for both within- and between-assay imprecision and the percentage inaccuracy for the quality control samples including lower and upper limits of quantitation were <12.1% and <11%, respectively. The absolute mean recovery of tetracaine was >97%. The accuracy and selectivity of this method allowed the rapid measurement of tetracaine in tape samples obtained from a skin tape stripping study of local anaesthetics in healthy subjects.

IL-21R is essential for epicutaneous sensitization and allergic skin inflammation in humans and mice

Atopic dermatitis (AD) is a common allergic inflammatory skin disease caused by a combination of intense pruritus, scratching, and epicutaneous (e.c.) sensitization with allergens. To explore the roles of IL-21 and IL-21 receptor (IL-21R) in AD, we examined skin lesions from patients with AD and used a mouse model of allergic skin inflammation. IL-21 and IL-21R expression was upregulated in acute skin lesions of AD patients and in mouse skin subjected to tape stripping, a surrogate for scratching. The importance of this finding was highlighted by the fact that both Il21r-/- mice and WT mice treated with soluble IL-21R-IgG2aFc fusion protein failed to develop skin inflammation after e.c. sensitization of tape-stripped skin. Adoptively transferred OVA-specific WT CD4+ T cells accumulated poorly in draining LNs (DLNs) of e.c. sensitized Il21r-/- mice. This was likely caused by both DC-intrinsic and nonintrinsic effects, because trafficking of skin DCs to DLNs was defective in Il21r-/- mice and, to a lesser extent, in WT mice reconstituted with Il21r-/- BM. More insight into this defect was provided by the observation that skin DCs from tape-stripped WT mice, but not Il21r-/- mice, upregulated CCR7 and migrated toward CCR7 ligands. Treatment of epidermal and dermal cells with IL-21 activated MMP2, which has been implicated in trafficking of skin DCs. These results suggest an important role for IL-21R in the mobilization of skin DCs to DLNs and the subsequent allergic response to e.c. introduced antigen.

Effects of lotioned disposable handkerchiefs on skin barrier recovery after tape stripping

In the present work, it was studied whether repeated use of lotioned disposable handkerchiefs on tape-stripped forearm skin was able to improve skin barrier recovery. Skin assessments included scoring of visual erythema and dryness/scaliness; and measuring of skin redness (Chromameter CR300), skin hydration (Corneometer CM825), and transepidermal water loss (Tewameter TM300). Four different lotioned paper handkerchiefs - randomly assigned to one of two subject groups (n=20) - were tested vs. the non-lotioned control handkerchief. The results were also compared with those obtained using a topically applied oil-in-water barrier cream (Dermalex). The three-day lasting protocol revealed that handkerchief wiping itself delayed skin recovery, but a significantly better performance was seen for the lotioned handkerchiefs containing fatty alcohols and mineral oils. This shows that the use of lotioned tissues helps to prevent skin damage inevitably caused by the wiping process. The controlled pre-damaged forearm method with tape stripping appears to be a suitable model to study the effects of repetitive wiping on irritated skin with disposable handkerchiefs of different quality. More specifically, the model seems applicable to mimic the nasolabial skin damage observed during a common cold associated with frequent use of disposable handkerchiefs.

The lymph vessel network in mouse skin visualised with antibodies against the hyaluronan receptor LYVE-1

Langerhans cells and dermal dendritic cells migrate to the draining lymph nodes through dermal lymphatic vessels. They do so in the steady-state and under inflammatory conditions. Peripheral T cell tolerance or T cell priming, respectively, are the consequences of migration. The nature of dendritic cell-containing vessels was mostly defined by electron microscopy or by their lack of blood endothelial markers. Selective markers for murine lymph endothelium were hitherto rare or not available. Here, we utilised recently developed antibodies against the murine hyaluronan receptor, LYVE-1, to study the lymph vessel network in mouse skin in more detail. In hairless skin from the ears, lymph vessels were spread out in a horizontal plane. They formed anastomoses, and they possessed frequent blind endings that were occasionally open. Lymph vessels were wider than blood vessels, which were identified by their strong CD31 expression. In body wall skin LYVE-1 reactive vessels did not extend laterally but they dived straight down into the deeper dermis. There, they are connected to each other and formed a network similar to ear skin. The number and width of lymph vessels did not grossly change upon inflammatory stimuli such as skin explant culture or tape stripping. There were also no marked changes in caliber in response to the TLR 7/8 ligand Imiquimod. Double-labelling experiments of cultured skin showed that most of the strongly cell surface MHC II-expressing (i.e. activated) dendritic cells were confined to the lymph vessels. Langerin/CD207 + cells within this population appeared later than dermal dendritic cells, i.e. langerin-negative cells. Comparable results were obtained after stimulating the skin in vivo with the TLR 7/8 ligand Imiquimod or by tape stripping. In untreated skin (i.e. steady state) a few MHC II + and Langerin/CD207 + cells, presumably migrating skin dendritic cells including epidermal Langerhans cells, were consistently observed within the lymph vessels. The novel antibody reagents may serve as important tools to further study the dendritic cell traffic in the skin under physiological conditions as well as in conditions of adoptive dendritic cell transfer in immunotherapy.

Development and validation of HPLC method for imiquimod determination in skin penetration studies

A simple, rapid and sensitive analytical procedure for the measurement of imiquimod in skin samples after in vitro penetration studies has been developed and validated. In vitro penetration studies were carried out in Franz diffusion cells with porcine skin. Tape stripping technique was used to separate the stratum corneum (SC) from the viable epidermis and dermis. Imiquimod was extracted from skin samples using a 7:3 (v/v) methanol:acetate buffer (100 mM, pH 4.0) solution and ultrasonication. Imiquimod was analyzed by HPLC using C(8) column and UV detection at 242 nm. The mobile phase used was acetonitrile:acetate buffer (pH 4.0, 100 mM):diethylamine (30:69.85:0.15, v/v) with flow rate 1 mL/min. Imiquimod eluted at 4.1 min and the running time was limited to 6.0 min. The procedure was linear across the following concentration ranges: 100-2500 ng/mL for both SC and tape-stripped skin and 20-800 ng/mL for receptor solution. Intra-day and inter-day accuracy and precision values were lower than 20% at the limit of quantitation. The recovery values ranged from 80 to 100%. The method is adequate to assay imiquimod from skin samples, enabling the determination of the cutaneous penetration profile of imiquimod by in vitro studies.

Lanolin-derived lipid mixtures mimic closely the lipid composition and organization of vernix caseosa lipids

The aim of the present study was to use semi-synthetic lipid mixtures to mimic the complex lipid composition, organization and thermotropic behaviour of vernix caseosa (VC) lipids. As VC shows multiple protecting and barrier supporting properties before and after birth, it is suggested that a VC substitute could be an innovative barrier cream for barrier deficient skin. Lanolin was selected as the source of the branched chain sterol esters and wax esters — the main lipid classes of VC. Different lipid fractions were isolated from lanolin and subsequently mixed with squalene, triglycerides, cholesterol, ceramides and fatty acids to generate semi-synthetic lipid mixtures that mimic the lipid composition of VC, as established by high-performance thin-layer chromatography. Differential scanning calorimetry and Fourier transform infrared spectroscopy investigations revealed that triglycerides play an important role in the (lateral) lipid organization and thermotropic behaviour of the synthetic lipid mixtures. Excellent resemblance of VC lipids was obtained when adding unsaturated triglycerides. Moreover, these lipid mixtures showed similar long range ordering as VC. The optimal lipid mixture was evaluated on tape-stripped hairless mouse skin in vivo. The rate of barrier recovery was increased and comparable to VC lipid treatment.

Expression of the L-fucose moiety on epidermal keratinocytes in psoriasis induced by the Koebner phenomenon: a sequential study.

The expression of Ulex europaeus agglutinin (UEA I) binding sites on cell-surface glycoproteins has been used as a marker for terminal differentiation. Increased number of UEA I binding sites of L-fucose specificity have been demonstrated in psoriatic epidermis. The results of lectin-binding studies in a series of biopsies taken sequentially (0 min, 5 min, 24 h, 7 days and 8 weeks) after tape-stripping of uninvolved skin in 12 psoriatic patients (three of whom were taking diltiazem, a calcium blocker at the time of the study) and six controls are presented. UEA I binding sites, which were expressed on the granular layer and upper layers of the stratum spinosum of pre-tape stripped uninvolved skin in psoriatic individuals, were progressively more numerous, with the expression of the L-fucose moiety on the lower stratum spinosum keratinocytes in the 7-day post-tape-stripping biopsies and 8-week biopsies, correlating with a moderate and marked increase in the proliferative index, respectively. In the Koebner-negative and non-psoriatic individuals who failed to develop psoriasis after tape-stripping, the UEA I binding sites were not expressed on keratinocytes of the lower stratum spinosum in any of the biopsies, although a mild increase in the proliferative index was noted in the 7-day biopsies. Our data suggest that the increased commitment of keratinocytes to terminally differentiate may be involved in the psoriatic process.(ABSTRACT TRUNCATED AT 250 WORDS)

Assessment of Quantum Dot Penetration into Intact, Tape-Stripped, Abraded and Flexed Rat Skin

Quantum dot (QD) nanoparticles have received attention due to their fluorescent characteristics and potential use in medical applications. Skin penetration is one of the major routes of exposure for nanoparticles to gain access to a biological system. QD655 and QD565 coated with carboxylic acid were studied for 8 and 24 h in flow-through diffusion cells with flexed, tape-stripped and abraded rat skin to determine if these mechanical actions could perturb the barrier and affect penetration. Nonflexed skin did not show QD penetration at 8 or 24 h. Flexed skin showed an increase in QD on the surface of skin but no penetration at 8 and 24 h. Tape-stripped skin depicted QD only on the surface of the viable epidermis. QD655 penetrated into the viable dermal layers of abraded skin at both 8 and 24 h, while QD565 was present only at 24 h. QD were not detected in the perfusate by fluorescence and inductively coupled plasma-optical emission spectroscopy analysis for cadmium at any time point. These results indicate that the rat skin penetration of QD655 and QD565 is primarily limited to the uppermost stratum corneum layers of intact skin. Barrier perturbation by tape stripping did not cause penetration, but abrasion allowed QD to penetrate deeper into the dermal layers.

Alterations in the desquamation-related proteolytic cleavage of corneodesmosin and other corneodesmosomal proteins in psoriatic lesional epidermis

Desquamation occurs after proteolysis of corneodesmosomal proteins, including corneodesmosin (CDSN), by proteases of the kallikrein family, particularly KLK7. Impaired desquamation is one of the features of psoriasis, and psoriasis-associated single nucleotide polymorphisms of the CDSN gene may potentially modify the proteolysis of the encoded protein. To test whether the proteolysis of CDSN and other corneodesmosomal components is altered in psoriatic epidermis. Total protein extracts obtained by tape-stripping of nonlesional and lesional skin from 11 patients were compared by immunoblotting experiments. An almost intact form of CDSN that has never been observed previously in the normal upper stratum corneum was detected in the lesional skin extracts, showing an altered proteolytic processing of the protein. This form was also observed in the nonlesional skin extracts, but in lower amounts. For most patients, increased amounts of desmoglein 1, plakoglobin and of high molecular weight fragments of desmocollin 1 were detected in the lesional skin. For most of them, similar amounts of KLK7 were immunodetected in both nonlesional and lesional skin extracts. No particular differences were observed related to the psoriasis type, the HLA-Cw6 status of the patients or any particular CDSN polymorphisms. We detected a near full-length form of CDSN that has not been previously observed in normal stratum corneum. The results suggest a reduced degradation of all corneodesmosomal proteins in psoriatic lesions which probably reflects the persistence of corneodesmosomes.

Mechanical injury by tape stripping causes Th2 responses in mice

Atopic dermatitis (AD) is a chronic inflammatory skin disease associated with cutaneous hyperreactivity to environmental allergens. In AD, skin injury caused by scratching and subsequent entry of antigen in the skin is critical to disease development. We hypothesize that DCs that emigrate from mechanically injured skin polarize the T cell response towards Th2. We used the hapten FITC as a marker to isolate DCs that have recently migrated from skin to draining lymph node (DLN). FITC was applied to shaved and tape‐stripped mouse skin, a surrogate for scratching, or to shaved skin without tape stripping. After 24 hrs, CD11cFITChi cells were enumerated in DLN, and examined for their capacity to cause proliferation and cytokine secretion in T cells. In each of the two groups CD11c+FITC+ constituted ~10% of DCs in DLNs and supported comparable proliferation of naïve DO11.10 CD4+T cells to OVA323–339 peptide. In contrast, FITC+ DCs from DLN of shaved/stripped skin induced significantly greater secretion of the Th2 cytokines IL‐4, IL‐5, IL‐13 and IL‐10, and less production of the Th1 cytokine IFN‐γ. IL‐10 is released in skin following mechanical injury. FITC+ DCs from DLN of IL‐10−/− mice were impaired in their ability to support Th2 cytokine secretion, consistent with our previous finding that the Th2 response to EC sensitization was impaired in these mice J Clin Invest. 114:399–407, 2004). The results demonstrate that mechanical skin injury polarizes skin DCs to support Th2 differentiation of naïve T cells; this is in part dependent on IL‐10.

Quantitative monitoring of dermal and inhalation exposure to 1,6-hexamethylene diisocyanate monomer and oligomers

Respiratory sensitization and occupational asthma are associated with exposure to 1,6-hexamethylene diisocyanate (HDI) in both monomeric and oligomeric forms. The monomer and polymers of diisocyanates differ significantly in their rates of absorption into tissue and their toxicity, and hence may differ in their contribution to sensitization. We have developed and evaluated a liquid chromatography/mass spectrometry (LC-MS) method capable of quantifying HDI and its oligomers (uretidone, biuret, and isocyanurate) in air, tape-stripped skin, and paint samples collected in the automotive refinishing industry. To generate analytical standards, urea derivatives of HDI, biuret, and isocyanurate were synthesized by reaction with 1-(2-methoxyphenyl)piperazine and purified. The urea derivatives were shown to degrade on average by less than 2% per week at -20 degrees C over a 2 month period in occupational samples. The average recovery of HDI and its oligomers from tape was 100% and the limits of detection were 2 and 8 fmol microl(-1), respectively. Exposure assessments were performed on 13 automotive spray painters to evaluate the LC-MS method and the sampling methods under field conditions. Isocyanurate was the most abundant component measured in paint tasks, with median air and skin concentrations of 2.4 mg m(-3) and 4.6 microg mm(-3), respectively. Log-transformed concentrations of HDI (r = 0.79, p < 0.0001) and of isocyanurate (r = 0.71, p < 0.0001) in the skin of workers were correlated with the log-transformed product of air concentration and painting time. The other polyisocyanates were detected on skin for less than 25% of the paint tasks. This LC-MS method provides a valuable tool to investigate inhalation and dermal exposures to specific polyisocyanates and to explore relative differences in the exposure pathways.

Involvement of organic anion transport system in transdermal absorption of flurbiprofen

We previously demonstrated that transdermal permeation of flurbiprofen is mediated by a nonlinear transport mechanism(s). Here, we aimed to characterize this transport mechanism by employing an Ussing-type chamber method with tape-stripped hairless mouse skin. Transdermal permeation of [ 3H]flurbipofen was vectorial, saturable and energy-dependent, suggesting the involvement of a carrier-mediated transport system. Transdermal permeation and uptake from the epidermal side of [ 3H]flurbiprofen were inhibited by various nonsteroidal anti-inflammatory drugs (NSAIDs). The inhibitory potency did not correlate well with lipophilicity; anionic NSAIDs tended to be more potent inhibitors than non-anionic NSAIDs. The inhibition profile of both [ 3H]flurbiprofen permeation and uptake, and the Michaelis constants, were similar for a given anionic compound. These results suggest that an organic anion transport system is involved in flurbiprofen uptake from the epidermal side during the process of transdermal absorption. Efflux of [ 3H]flurbiprofen from the skin to the epidermal side, but not to the hypodermal side, increased in the presence of flurbiprofen or several anionic compounds. Such trans-stimulation may suggest the involvement of an organic anion exchanger system. Organic anion transporter 2 (OAT2) is a candidate for the exchanger involved in uptake and/or efflux of flurbiprofen in the skin.

Ceramide analogue 14S24 selectively recovers perturbed human skin barrier

Topical ceramide application is an effective therapeutic approach in skin disorders with disturbed barrier function, including atopic dermatitis and psoriasis. To evaluate ceramide analogue N-tetracosanoyl-(l)-serine tetradecyl ester (14S24) using a novel ex vivo model. Freshly excised human skin was disrupted by lipid extraction, tape stripping and sodium lauryl sulphate (SLS) treatment. Barrier perturbation was evaluated by the measurement of transepidermal water loss (TEWL), skin hydration and the penetration of model compound, theophylline (TH), assessed by microdialysis. The effect of topical 5% 14S24 was compared with a commercial formulation containing a skin lipid mixture (LR) and control formulation with no skin lipids (L). Both LR and 14S24 produced significant recovery of TEWL and TH penetration in extracted and tape-stripped skin with 14S24 being significantly more effective. In SLS-treated skin, 14S24 decreased TEWL but not TH penetration; LR was inactive. L improved skin hydration but not barrier characteristics. Weak correlation between TEWL and TH penetration was observed in extracted and tape-stripped skin but not in SLS-treated skin. Cutaneous microdialysis can serve as a useful tool for the evaluation of skin barrier recovery by topical formulations ex vivo whereas TEWL may not be an appropriate measure of skin barrier function in such studies. The excellent barrier repair activity of 14S24 could be beneficial in skin disorders with ceramide deficiency.

Smad3 Signal Transducer Regulates Skin Inflammation and Specific IgE Response in Murine Model of Atopic Dermatitis

Atopic dermatitis (AD) is a common chronic inflammatory skin disease characterized by itchy, dry, and inflamed skin. Transforming growth factor (TGF)-beta is an important fibrogenic and immunomodulatory factor that regulates cellular processes in the injured and inflamed skin. This study examines the role of the TGF-beta-Smad signaling pathway using Smad3-deficient mice in a murine model of AD. Dermatitis was induced in mice by epicutaneous application of ovalbumin (OVA) applied in a patch to tape-stripped skin. OVA-specific IgE and IgG2a antibody levels were measured by ELISA. Skin biopsies from sensitized skin areas were used for RNA isolation, histology, and immunohistochemical examination. The thickness of dermis was significantly reduced in OVA-sensitized skin of Smad3-/- mice. The defect in the dermal thickness was accompanied by a decrease in the expression of mRNA for proinflammatory cytokines IL-6 and IL-1beta in the OVA-sensitized skin. In contrast, the number of mast cells was significantly increased in OVA-sensitized skin of Smad3-/- mice, which also exhibited elevated levels of OVA-specific IgE. These results demonstrate that the Smad3-pathway regulates allergen-induced skin inflammation and systemic IgE antibody production in a murine model AD. The Smad3 signaling pathway might be a potential target in the therapy of allergic skin diseases.

Validation and application of capillary electrophoresis for the analysis of lidocaine in a skin tape stripping study

A fast and simple capillary zone electrophoresis method was developed and validated for the determination of lidocaine in skin using tape samples. Separation was performed in a 350 mm (265 mm to window) x 50 microm i.d. fused silica capillary using a background electrolyte of phosphoric acid-Tris pH 2.5. The extraction of lidocaine from tape samples was achieved using methanol, which was diluted to 50% with water before injection. Procaine was the internal standard. The migration times for procaine and lidocaine were 2.9 and 3.2 min, respectively. The limit of quantification for lidocaine was 50 microg, with signal to noise ratio greater than 10. The calibration curve was linear from 50 to 1000 microg with r(2) greater than 0.99. The CV for both within- and between-assay imprecision and the percentage of inaccuracy for the quality control samples including lower and upper limits of quantitation were <or=2% and <or=14%, respectively. The absolute recovery of lidocaine was >97%. The accuracy and selectivity of this method allowed the measurement of lidocaine in tape samples obtained from a skin tape stripping study of local anesthetics in healthy subjects.

Evaluation of skin penetration of topically applied drugs in humans by cutaneous microdialysis: acyclovir vs. salicylic acid

Cutaneous drug application is used for both local drug therapy and systemic treatment. For both types of treatment, the drug concentration profile in, and transport across, the skin is important. To evaluate skin penetration of topically-applied drugs we recently used cutaneous microdialysis. The aim of this study was the use of this method for studying acyclovir and salicylic acid. Five per cent acyclovir cream was applied on intact and tape-stripped skin of healthy volunteers and 5% salicylic acid ointment-onto intact skin of other volunteers. Microdialysis probes with 2 kDa molecular weight cut-off were inserted intradermally and were perfused with Ringer solution. Drug concentrations were measured by high-performance liquid chromatography. Following topical application of 5% acyclovir cream onto intact skin of eight healthy volunteers, no drug was determinable in the skin (cutaneous microdialysate) in any of the subjects studied. After partial removal of the stratum corneum the penetration of this drug into skin increased markedly. The mean maximum skin concentration was about 2 x 5 micromol/L after 2 x 4 +/- 0 x 7 h. Topically applied salicylic acid penetrated intact skin with a maximum concentration in the cutaneous microdialysate of 7 x 57 +/- 3 x 90 micromol/L after 5 x 3 +/- 0 x 4 h. Cutaneous microdialysis is a valuable method for estimating skin concentration of topically-applied drug. It allows evaluation after application to a small skin area, of about 2 cm(2), thereby reducing the risk of systemic toxicity. The method may be helpful for evaluating the influence of skin condition on the transport process.

Cyclooxygenase-2 Inhibition Promotes Enhancement of Antitumor Responses by Transcutaneous Vaccination with Cytosine-Phosphate-Guanosine-Oligodeoxynucleotides and Model Tumor Antigen

One of the principal goals in tumor immune prophylaxis and tumor therapy is the induction of antitumor responses by generating sufficient numbers of tumor antigen-specific helper T (Th)1 cells and cytotoxic T lymphocytes (CTLs). We have demonstrated that the administration of cytosine-phosphate-guanosine-oligodeoxynucleotide (CpG-ODN) through tape-stripped skin induced a Th1-type immune response and suggested that the skin is a potential site for vaccination. CpG-ODN induces the expression of cyclooxygenase (COX)-2, and its product prostaglandin (PG) E2 underlies an immunosuppressive network, therefore it is a simple strategy to use a COX-2 inhibitor for tumor vaccination with CpG-ODN. In this study, we examined whether a COX-2 inhibitor enhances the antitumor immune response induced by CpG-ODN with model tumor antigen, ovalbumin (OVA), applied to tape-stripped skin in mice. The COX-2 inhibitor remarkably enhanced antigen-specific Th1-type immune responses and generation of CTLs induced by transcutaneous vaccination with CpG-ODN and OVA. PGE2 and IL-10 levels in the skin were significantly decreased and production of IL-12 was enhanced. This vaccination also induces an effective antitumor immunity in tumor-challenged mice. These results suggested that transcutaneous vaccination with a COX-2 inhibitor, CpG-ODN, and tumor antigen is a very simple and cost-effective strategy for tumor vaccine and may be readily achievable.

Toll-like receptor-9 expression induced by tape-stripping triggers on effective immune response with CpG-oligodeoxynucleotides

Recently, there has been a lot of interest in the potential of non-invasive routes, such as via the skin, for vaccine delivery. CpG-oligodeoxynucleotides (ODN) are an effective adjuvant for the induction of cellular and humoral immunities when administered with an antigen. We demonstrated here that tape-stripping induces the expression of toll-like receptor (TLR)-9 in the skin, and enhances the Th1-type immune response triggered by CpG-ODN administered through the tape-stripped skin. Tape-stripping induces expression of TLR-9 and tumor necrosis factor (TNF)-α in the skin, and CpG-ODN treatment through the tape-stripped skin enhances the migration of antigen presenting cells (APCs) to the draining lymph nodes. On the other hand, TLR-9 mRNA and TNF-α mRNA were not observed in the skin when CpG-ODN was injected intradermally in a volume of 10 μL, or in a Th1-type immune response. The transdermal application of CpG-ODN with an antigen through the tape-stripped skin is an effective way to induce a Th1-type immune response, and is also a simple, cost-effective and needle-free vaccination system.

Trace analysis of fullerenes in biological samples by simplified liquid–liquid extraction and high-performance liquid chromatography

Fullerene (C 60) has several potential biomedical and industrial applications. While pure fullerene is not soluble in water, nanoparticles of the fullerene aggregates (nano-C 60) can be prepared in water solutions. The concentration of nano-C 60 in biological media after systemic exposure could be very low and requires trace analytical methods to be developed for the toxicological and pharmacokinetic studies of the nanomaterial. A serious drop in extraction efficiency was observed when the concentration was under 0.5 μg/mL using traditional liquid–liquid extraction (LLE) protocols. The evaporation of the solvent extract to dryness was found to be the main reason for the efficiency drop and an improved evaporation method was proposed to overcome this problem. Optimal proportion of glacial acetic acid (GAA) was used to solublize the proteins and surfactants in the biological samples, so that the emulsion problem was eliminated during LLE. Magnesium perchlorate was used to destabilize the nano-C 60 particles in the water solution and promoted the solvent extraction. A simplified LLE method was developed for high throughput while preserved the advantages of the traditional LLE. The developed method was used for trace analysis of fullerenes in protein containing media and tape-stripped skin samples. Under optimal experimental conditions, the detection limit was 0.34 ng/mL and the recovery was in the range of 94–100% ( n = 5) at a concentration of 10 ng/mL nano-C 60 in the biological media.

Regional differences in adhesive tape stripping of human skin

The regional differences in acrylic adhesive tape stripping of human skin were evaluated. Two kinds of adhesive tape with different water vapor permeabilities were applied to the flexor side of the forearm and the cheek, palm and sole in eight healthy male volunteers. Dermal peeling force was measured at 1 and 6 h after application. Simultaneously, the electrical conductance of the skin beneath the applied tape and the amount of stripped corneocytes on the removed tape were measured. The regional difference in dermal peeling force of the permeable tape was large and that of the occlusive tape was small. The dermal peeling force of the permeable tape was high in the order of the palm, sole, cheek and forearm and individual differences among subjects in dermal peeling force of tape applied to the palm was large. The regional difference in dermal peeling force was considered to be related to fluid accumulation beneath the applied tape and the amount of stripped corneocytes. On the other hand, that the dermal peeling force of the occlusive tape was low and the regional difference was small may have been caused by fluid accumulation from the skin beneath the applied tape on all regions.