Fluorescence enhanced BA-LIFT for single cell detection and isolation.

Laser bioprinting is a term that refers to a group of laser-based techniques for printing living cells with high precision and good viability. Most of these techniques are based on modifications of the standard laser induced forward transfer technique (LIFT). When it comes to printing living material, direct laser irradiation should be avoided, therefore an indirect LIFT technique comprising an energy absorption layer should be used. This work presents a blister actuated-LIFT (BA-LIFT) technique which uses a commercial polyimide tape as a uniform energy absorption layer. To increase the potential of the technique for cell selection and printing accuracy, we take advantage of the high optical transmission of the polyimide layer to implement an in-line fluorescence and conventional imaging vision system coaxial with the laser path. With this system and using the appropriate staining methodology it is possible to track and identify different cell types, selecting those to be transferred and tracking cell survival both in the donor and acceptor substrates. We studied the BA-LIFT printability map for sodium alginate and methylcellulose hydrogels in the fluence range from 6.1 to 2.0 J cm-2 together with the cell viability assessment measured by our fluorescence system. The study has revealed that less concentrated, therefore less viscose hydrogel shows better results with lower fluences, whereas hydrogels with higher concentrations present better results at higher fluences. Also, at low fluences 98 % of cell viability was obtained, besides both primary cells and cell lines keep their integrity, proliferating and functional activity. The technique was tested by tracking and targeting mouse hematopoietic progenitor stem cells transferred to assess colony forming units; moreover, natural killer cells were isolated and its activation in a stimulation media was tracked with the fluorescence system.

Effect of lairage time ( 0h, 3 h, 6 h or 12 h) on glycogen content and meat quality parameters in suckling lambs.

The effect of slaughter at arrival (L0) or after 3 (L3), 6 (L6), 12 (L12) h of lairage on carcass and meat quality of suckling lambs has been studied. Lairage time had a slight effect on carcass quality traits, but most of the meat quality parameters at 24h post mortem were affected. Weight losses increased and glycogen content of liver and longissimus muscle decreased as lairage time increased. Longissimus muscle from L3 lambs had the highest pH, shear force and toughness and the lowest b* and C* values, at 24h post mortem. L3 and L6 had the higher proportion of expelled juice, or low water holding capacity (WHC), at 24h post mortem. Texture parameters and WHC were similar among groups after 5 days of storage. Despite the fact that the effects of lairage time on meat quality disappears after storage, from the point of view of weight losses the slaughter of suckling lambs at arrival is recommended.

Physiological response and carcass and meat quality of suckling lambs in relation to transport time and stocking density during transport by road.

To evaluate the effect of stocking density and transport time on physiological responses and meat quality, 72 male suckling lambs were transported by road to the slaughterhouse at three different stocking densities (0.12, 0.20 or 0.25 m2/lamb) and two transport times (5 h or 30 min). Blood samples were collected pre-transport at the farm and after unloading in the slaughterhouse to measure levels of cortisol, creatine kinase (CK) and lactate dehydrogenase (LDH). After slaughter, the weights of the hot carcass, liver and spleen were recorded and pH in Musculus longisimus thoracis et lumborum (L), Musculus semitendinosus (ST) and Musculus psoas major (PM) were determined. Colour, water-holding capacity (WHC), texture and thiobarbituric acid-reactive substances (TBARS) values were measured in samples of L, at 24 h post mortem and after 5 days of ageing. Cortisol and LDH were higher in suckling lambs transported for 30 min than those transported for 5 h. Stocking density did not affect blood parameters studied. Transport time significantly affected some carcass quality parameters, but stocking density had no significant effect. Suckling lambs transported for 5 h had lower liver weights and dressing percentages than those transported for 30 min. Transport time influenced pH values, with lambs subjected to the longer journey showing the lowest pH at 0 h in the three muscles studied, with the lowest final pH in L and PM. The PM lambs transported at high density (0.12 m2/lamb) had the lowest pH at 24 h. Transport time and stocking density did not greatly affect colour and texture parameters. The meat from lambs transported for 30 min had higher WHC than meat from lambs transported for 5 h. Animals transported for longer journeys showed higher lipid oxidation after 5 days of ageing than those transported for 30 min. Loading and initial transport caused significant stress response in suckling lambs, that stress response was reduced over the time course of the journey.

Estimation of α-tocopherol concentration necessary to optimise lamb meat quality stability during storage in high-oxygen modified atmosphere using broken-line regression analysis.

The research was carried out to evaluate the effect of different α-tocopherol concentrations in lamb meat on oxidative stability during storage in high-oxygen atmosphere. Thirty-six lambs were randomly distributed to four groups and given diets containing four levels of vitamin E (20, 270, 520 and 1020 mg vitamin E/kg feed) from an initial weight of 13.2 ± 0.5 kg to a slaughter weight of 26.2 ± 0.3 kg. Supplementation of the diet with vitamin E increased (P < 0.001) the concentration of α-tocopherol in the meat and concentrations were obtained in the 0.46 to 4.14 mg/kg meat range. Broken-line analysis of data indicated a target dietary vitamin E supplementation of 287 mg/kg feed, which corresponded with a concentration of 2.26 mg α-tocopherol/kg meat. α-Tocopherol in meat was highly correlated with the oxidation of lipids and pigments. Broken-line analysis of data indicated the target α-tocopherol concentration in lamb for improved protection against lipid and pigment oxidation during 14, 21 and 28 days of storage in high-oxygen atmosphere was in the range 1.87 to 2.37 mg/kg meat. These concentrations of α-tocopherol in the meat made it possible to maintain the indicator values of lipid and pigment oxidation below the values considered in the bibliography as unacceptable to the consumer.

The influences of carcass weight and depot on the fatty acid composition of fats of suckling Manchego lambs.

The effects of carcass weight (<5.5 kg, 5.5-6.5 kg, ⩾6.5 kg) of Manchego suckling lambs, type of fat depot of the leg (subcutaneous, intermuscular and intramuscular) and muscle (m. longissimus dorsi (LD) or m. quadriceps femoris (QF)) on the fatty acid composition were studied. Carcass fatness increased with increasing carcass weight. However, few differences in fatty acid proportion were observed between the three carcass weight groups. The lightest carcasses had lower proportions of myristic (C14:0) and palmitoleic (C16:1) acid and higher proportions of stearic (C18:0) acid, desirable fatty acids (DFA) (C18:0+total unsaturated fatty acid (TUFA)) and a greater nutritive value ((C18:0+C18:1)/C16:0). The fatty acid composition of subcutaneous and intermuscular fat depots of the leg were similar, whereas the intramuscular fat depot was different from both of these, since it had a lower proportion of saturated fatty acids (SFA), a greater proportion of TUFA, higher PUFA/SFA and n-6/n-3 ratios, and greater DFA and nutritive values. The intramuscular fat of the LD was more saturated than QF, higher SFA, and had lower value of PUFA/SFA ratio and DFA. These results indicated that the intramuscular fat quality of the leg was better than subcutaneous and intermuscular fat depots and also that intramuscular fat displayed better attributes of quality (lower SFA, higher TUFA, and greater PUFA/SFA ratio and DFA as well) than intramuscular fat of the LD from the consumer health standpoint. Despite carcass weight affected carcass fatness, it did not seem to influence the proportions of saturated or polyunsaturated fatty acids. For this reason, carcass weight probably had little effect on the organoleptic characteristics of the meat.

Prediction of suckling lamb carcass composition from objective and subjective carcass measurements.

Correlations and regression between fatness and carcass conformation measurements and carcass tissue composition has been studied for both sexes in suckling lambs, with slaughter weights between 9 and 15 kg. Objective fat measurements showed higher correlations than subjective fat measurements, mainly with carcass fat. Conformation measurements presented greater correlations with tissues weight than with tissues proportion (r⩽0.50). CCW was highly correlated with total muscle in the carcass (R(2)=0.961). Two fatness measurements (KKCF proportion and dorsal fat thickness) were included for the prediction of the muscle proportion. Furthermore, the model included other measurements, such as fore cannon bone weight and internal carcass length although they displayed very low correlation coefficients. This equation explained 64.1% of the variation of carcass muscle proportion (r.s.d.=1.532). CCW was included in the prediction equations of tissue weights, although in the fat tissue equation it appeared in the form of the carcass compactness index (CCW/L). CCW was a good predictor of the weight of carcass tissues in suckling lambs, especially for muscle. Prediction equations for tissue composition in grams were more accurate (R(2)⩾0.91) than those for tissue proportion. When gender was included in all models it did not improve either accuracy or precision of the prediction.

Carcass and meat quality of light lambs using principal component analysis.

Eighty-six male light lambs of Manchego breed were used in this study. Principal component (PC) analysis was performed to study the relationship between carcass quality variables (n=22) and between meat quality measures (n=21). The carcass quality was assessed using objective and subjective measurements of conformation and fatness besides the joints proportion and tissues proportion of the leg. The measurements used to evaluate meat quality were pH in longissimus dorsi and semitendinosus muscles, the colour, moisture, water holding capacity, cooking losses, texture and sensorial analysis on longissimus dorsi. The five first PCs explained about 77% of the total variability for carcass measures whereas for meat quality the 74% of the total variability was explained for the eight first PCs. All the carcass measurements showed similar weight to define the first PC, whereas the muscle and bone proportion as well as muscle:bone ratio of the leg were useful to define the second PC. The meat quality measures that were more effective to define the first PC were the meat colour measurements, whereas the sensorial variables defined the second PC. The projection of the carcass quality data in the first two PCs allowed to distinguish clearly between heavier carcasses (higher than 6.5 kg) and lighter carcasses (lower than 5.5 kg). The carcasses with a weight higher than 6.5 kg were on the left side of the figure, where the variables of conformation and fatness lie. The group of medium carcass weight were placed between the two previous groups. The projection of the meat quality data in the first two PCs did not differ between hot carcass weights, although there was a trend, the lighter carcasses lay on the left side of the graph, which implies small differences between meat quality in this range of carcass weight.

Carcass classification in suckling lambs. II. Comparison among subjective carcass classification methods: fatness scales and conformation scales with 0.25 point-intervals.

Forty-nine Manchega breed suckling lambs were used. Carcass weight was less than 13 kg, and lambs were slaughtered at 10, 12 and 14 kg liveweight. Carcasses were classified by three assessors on colour photographs, by several classification methods: the European Union scale for light lambs carcasses (EU), Colomer-Rocher methods for fatness (CF) and conformation (CC), and the new scale for suckling lambs (SL) developed by our research group. Carcasses were also assessed using these scales divided into 0.25 point-intervals (EU-25, CF-25, CC-25 and SL-25). A correlation analysis was made between assessors score and tissue composition in the carcass and in three joints (leg, best end of neck and loin, and shoulder). For all the methods studied 0.25 point scales were best correlated to tissue composition than 1.0 points-interval scales. When 1.0 point-interval scales were used the SL scale was best correlated to tissue composition. When 0.25 point-scales were used, the EU scale gave the best correlations. Conformation scale (CC) was useful to predict carcass composition, specially kidney knob and channel fat content.

Effect of different feeds on meat quality and fatty acid composition of lambs fattened at pasture.

Two kinds of feed (commercial concentrate vs whole supplemented barley) were compared in unweaned lambs and lambs weaned at 40 days of age, fattened at pasture and slaughtered at 28 kg live weight, in order to observe their effects on meat quality and fatty acid composition. The weaning status influenced fatness; unweaned lambs displayed a greater carcass fatness score and more kidney knob and channel fat than weaned lambs. Compared with the unweaned animals, weaned lambs exhibited higher pH values at 0 h and 45 min in the m. longissimus thoracis (LT) and at 45 min and 24 h in the m. semitendinosus. The redness index (a*) of the m. LT of weaned lambs was higher than that of unweaned lambs, and lambs fed concentrate displayed a higher yellowness index (b*) and a higher Hue value than those given whole barley. Water-holding capacity did not vary with the treatments studied. The proportion of saturated fatty acids (SFA) in intramuscular fat was higher (P⩾0.001) in unweaned lambs than in weaned ones while, on the other hand, the latter displayed a higher proportion of monounsaturated fatty acids (MUFA) and a higher n-6/n-3 ratio in the same tissue. As was the case with intramuscular fat, the subcutaneous fat of unweaned lambs exhibited higher proportions of medium-chain fatty acids (C12:0, C14:0 y C16:0) and lower ones of stearic (C18:0) and oleic (C18:1) fatty acids than that of weaned lambs. Higher levels of heptadecenoic acid (C17:1) were found in the subcutaneous fat of lambs fed whole barley than in that of lambs given concentrate. PUFA/SFA and n-6/n-3 ratios were lower in the m. LT than in the m. quadriceps femoris.

Carcass classification in suckling lambs. Discrimination ability of the European Union scale.

Forty-nine Manchega suckling lambs were used in this trial. Lambs were slaughtered at 10, 12 and 14 kg liveweight (cold carcass weight lower than 13 kg). The degree of fatness was assessed by three assessors on colour photographs of the carcasses, using the European Union scale for light lambs (EU), and another new scale developed in our laboratory (Suckling Lamb scale, SL). Carcass degree of fatness was reassessed using the same scales divided into 0.25 point-intervals (EUI, SLI). Carcasses were allocated into four groups by means of a tree classification algorithm (cluster analysis), according to 27 variates, namely muscle percentage, whole fat percentage, subcutaneous fat, intermuscular fat, kidney knob and channel fat percentage and muscle percentage, both in the carcass and in three joints (leg, best end neck plus loin, and shoulder). SL scale groups showed a higher number of variates well correlated to assessors' scoring than EU scale groups. Nevertheless, the EU scale showed significant between-groups differences in a higher number of variates (P<0.01). Using 0.25-points interval scales, EUI scale scoring was the best correlated to tissue composition. However, most variates were better correlated to dorsal fat thickness measurement (4 cm back from last rib and 4 cm from the carcass midline) than to assessors' scoring. All four groups in the tree classification were statistically different from one another, but assessors only were able to detect differences between whole fat percentage in group 1 and the rest. These results suggest that it is possible to improve the photographic standards of the EU method by using a 0.25 points interval scale. In abattoir conditions, the new SL method (1.0 point intervals) proved to be better. Measure of dorsal fat thickness proved to be a very good predictor of carcass tissue composition, but it is a more expensive measure.

Physico-chemical characteristics of carcass and meat Manchego-breed suckling lambs slaughtered at different weights.

Forty-nine Manchego-breed lambs raised exclusively on their dams' milk until slaughter were used in this study. The effects of gender and slaughter weight (10, 12 and 14 kg) on carcass fatness, meat quality and the fatty acid composition of their fat were studied. Fatness, and in particular dorsal-fat thickness (P⩽0.01), increased with live weight. The effect of gender was even greater (P⩽0.001), as female lambs presented the highest fatness values for all parameters studied. The smallest drop in m. longissimus pH values was seen in the lowest-weight (10 kg) lambs. These same lambs displayed the highest L(∗) value and thus the lightest colour. Fatty acid composition, which was not influenced by live weight, was affected by gender. The subcutaneous fat of female lambs contained more linolenic acid (C18:3) (P⩽0.01) and a greater proportion of polyunsaturated fatty acids than that of male lambs (P⩽0.001). Likewise, the intramuscular fat of female lambs displayed a greater proportion of monounsaturated fatty acids (P⩽0.001) than that of male lambs.

Physico-chemical characteristics of carcass and meat Manchego-breed suckling lambs slaughtered at different weights.

Forty-nine Manchego-breed lambs raised exclusively on their dams' milk until slaughter were used in this study. The effects of gender and slaughter weight (10, 12 and 14 kg) on carcass fatness, meat quality and the fatty acid composition of their fat were studied. Fatness, and in particular dorsal-fat thickness (P⩽0.01), increased with live weight. The effect of gender was even greater (P⩽0.001), as female lambs presented the highest fatness values for all parameters studied. The smallest drop in m. longissimus pH values was seen in the lowest-weight (10 kg) lambs. These same lambs displayed the highest L* value and thus the lightest colour. Fatty acid composition, which was not influenced by live weight, was affected by gender. The subcutaneous fat of female lambs contained more linolenic acid (C18:3) (P⩽0.01) and a greater proportion of polyunsaturated fatty acids than that of male lambs (P⩽0.001). Likewise, the intramuscular fat of female lambs displayed a greater proportion of monounsaturated fatty acids (P⩽0.001) than that of male lambs.

Fatty acid composition of adipose depots of suckling lambs raised under different production systems.

Forty-nine male Talaverana-breed suckling lambs slaughtered at 14 kg live weight were used to study the fatty acid composition of adipose depots of lambs raised under different management systems (pasture or drylot) and diets (maternal milk with or without a concentrate supplement). Drylot lambs displayed a higher fatness score, in addition to greater dorsal fat thickness; these parameters corresponded with a greater percentage of total fat at dissection. Addition of concentrate to the diet did not modify carcass fatness. Drylot lambs presented higher proportions of oleic acid (C18:1) and monounsaturated fatty acids (MUFA) and lower linolenic acid (C18:3) values in subcutaneous and intramuscular fat (muscle longissimus thoracis). The subcutaneous fat of these lambs thus displayed a higher n-6/n-3 PUFA ratio (C18:2/C18:3) than that of pasture-raised ones. Concentrate in the diet of both pasture and drylot lambs resulted in higher linoleic fatty acid (C18:2) and PUFA values and a greater PUFA/SFA ratio in intramuscular fat than the corresponding values of lambs which consumed maternal milk alone. Loin displayed a higher percentage of subcutaneous and intramuscular chemical fat than leg. Compared with intramuscular leg fat (muscle quadriceps femoris), intramuscular loin fat (muscle longissimus thoracis) had lower oleic acid (C18:1) and MUFA values, higher PUFA values and a higher PUFA/SFA ratio, due to greater linolenic acid (C18:3) levels.